Good morning. I hope everyone had a good weekend. Today we are going to talk about three related topics, synaptic inhibition. We've not discussed that at all, cable properties of neurons, and then putting it all together, electrical integration inhibition cable properties. We'll use as an example the cerebellum. Here are the readings, and you'll also see that many of the illustrations in today's talk come from Candela, and so you can peruse those chapters as well. We're going to talk about the pentameric, GABA, and glycine receptors. As we'll see, they look a lot like acetylcholine receptors at the nerve muscle synapse. As we've said over and over again, the nerve muscle synapse is a model for all sorts of aspects of synaptic transmission, both function and development. Indeed, as we'll see, the GABA and glycine receptors are not cation channels like acetylcholine receptors. They are permeable to anions, mostly chloride of course, which is the dominant anion, negatively charged anion in the body. They are in fact so close to the structure of acetylcholine receptors that you can change a cation channel into an anion channel with only one amino acid change per subunit. Remember, the story about the five Caltech professors with their knees in the middle of the channel, they all apparently rotate their knees or straighten them up simultaneously, and yet the anion going through the channel can only be a cation ion, or in the case of a GABA receptor, it can only be an anion. Changing one of those amino acids very near the knee does the trick, though the selectivity is not so good as what a real GABA or glycine receptor would do. So GABA, gamma amino butyric acid, is the major inhibitory transmitter in the brain. Glycine is the dominant inhibitory transmitter in the spinal cord and in the hindbrain. GABA, actually GABA A receptors are the ones that look like nicotinic acetylcholine receptors. There are also G protein coupled receptors that are activated by GABA, just as there are G protein coupled receptors that are activated by acetylcholine. The GABA A receptors are more variable than the glycine receptors in subunit composition and therefore in kinetic behavior. As far as we know there are no G protein coupled glycine receptors among vertebrates or among invertebrates. Now I had to talk by analogy with nicotinic receptors until about a year and a half ago, at which point the crystal structure of a GABA A receptor was published. Very exciting observation and you've seen enough pictures of nicotinic acetylcholine receptors to know, not surprisingly, that at this level of resolution, a GABA A receptor looks pretty much like an acetylcholine receptor. It's got an extracellular portion which consists mostly of beta strands where the ligand, where the transmitter binds. I'll show you some more views of that in a moment. It's got a transmembrane domain, mostly consisting of alpha helices. And then it's got an intracellular, so here we have out and in, it's got an intracellular part which is not really very well resolved in most of the x-ray crystallographic structures, just like unfortunately for nicotinic receptors. And in fact here it's been cut off to make a construct that will crystallize. Now if we, if instead of looking from the side of the membrane, we look from the top of the membrane, we see the usual pentameric structure. We see the transmembrane domain lining what appears to be an ion channel. And we see, well this part is not so important. So again it's a homomer, this particular one is a homomer of five GABA A subunits. Remember I told you there were lots of different kinds of GABA A subunits. This one is the GABA A beta 3 subunit. And so it is quite gratifying to see this sort of structure. Now you remember we also said when we talked about nerve muscle synapses that an excitatory a synapse pushes the membrane potential toward the reversal potential called eREV for the synaptic channels. And so we get a push more positive to the threshold for canion channels, more negative with a reversal potential of around zero. So we would expect and it is indeed true that at GABA and glycine receptors which are permeant to chloride, we push the membrane potential toward the Nernst potential for chloride, which is pretty near the resting potential in most cells around minus 70 millivolts. Remember we did say during the lecture and development that in some immature cells the reversal potential is a little more positive. We won't go into that now. So here we have a push toward the resting potential, perhaps a little more hyper polarized than the resting potential. And so this would prevent a cell from firing spikes and there's a great deal of analysis of the way excitatory and inhibitory influences some on a neuron. Any questions so far? Okay. I'll give you a little sampling of what you'll learn about for neuro pharmacology in January of 2017 when I teach by 155. But there is a very rich and useful pharmacology of GABA receptors. There are, first of all, the activators. GABA is itself an activator and agonist from the Greek word to act. It's the natural agonist of GABA receptors. And just like acetylcholine and nicotine acetylcholine receptors, GABA actually binds at the interface between two subunits. How that actually causes the gate to open, we're not sure yet, but it does so. In addition, there is a very interesting large class of molecules that binds at the interface between other subunits. So this particular GABA receptor is a hetero pentamer to alpha one subunits to beta two subunits and one gamma two subunits. Remember, we said that the GABA receptors have all sorts of subunits and have varied kinetics and varied localization. Well, Valium, also called diazepam, binds not at the interface where the transmitter binds, but at another interface. And it is, you may have learned, and by eight by nine, about allosteric modulators. You certainly will learn this in biochem 110. So Valium is an allosteric modulator diazepam, sorry, is an allosteric modulator of the GABA receptor. There are many allosteric modulators of GABA receptors. Some of the trademarks are Ambien and Lunista. As usual, we don't remember trademarks in this course because there are various types of GABA receptors. Some of the diazepam-like molecules decrease anxiety. Others are pretty good sleeping pills and others have more of an sedative effect. So there's a rich field of ligands of molecules that bind to GABA receptors at the allosteric site. And this, what an allosteric modulator does is not to open the channel by itself, but to help the natural ligand GABA open the channel. Also, phenobarbital binds to GABA receptors and activates them. It's not certain yet where phenobarbital binds. There are also blockers for GABA and glycine receptors. They can bind in one of two places. Again, you'll learn more about this in by 155. Some of them can bind at the agonist site, at the GABA site, and prevent it from binding. Those would be competitive antagonists. A good example is called bicuculine. And as at other ion channels, there's a class of blockers that binds like a cork in a drain or in a wine bottle. And among those blockers are picrotoxin, which seems to sit right inside the channel and prevent conduction. Those GABA A blockers are relatively toxic drugs. The good example, for instance, about picrotoxin is that it causes spasms and convulsions. So they're not very useful clinically, but they are useful for research, for proving what sort of receptor or channel we have, or for emphasizing excitation over inhibition. Any questions? And again, we discussed the controversies about how the receptor transduces binding into channel gating, whether it is those professors rotating their knees or straightening their legs. Not yet clear, both ideas are also in play for GABA A and glycine receptors. The main problem is that we don't have x-ray crystallographic structures for a given receptor in both the open and the closed state. But it's really remarkable that we do have these structures in the first place. So we have pretty much completed our survey of synaptic receptors. We've talked about the family that includes acetylcholine, serotonin, and GABA. There are other members of the family, which we mentioned, for instance, the day the Nobel Prize for Ivermectin was announced. We noted that there are invertebrate glucial channels that are allosterically activated by Ivermectin, and then there are some other invertebrate channels. We also talked about most glutamate receptor channels, which have a totally different topology, a totally different number of subunits, except for the invertebrate glucial channels. And then we mentioned that there is a third class of synaptic receptors, synaptic receptors for ATP, and we're not going to have a chance to discuss those in any kind of detail in this course. They have a different membrane topology per subunit again, and they also have only three subunits per receptor. Quite interesting molecules. Now we begin to put together our knowledge about individual molecules, and we start thinking about how a neuron works. And so here in a figure that's very much like the one in Candel, but rotated, we talk about parts of two generalized central nervous system neurons, the presynaptic neuron and the post-synaptic neuron. And all of the parts are here. We have the cell, the, first of all, the inputs, the dendrites, dendrites comes from the Greek word for tree, and the apical dendrites are the furthest away there at the apex of the cell. We also have the basal dendrites that are nearest to the cell body. We have the initial segment of the axon, which is where the action potential is mostly generated. That's called the axon hillock, because it's a little wider, a little hill. We have the cell body itself. That's the soma. We have excitatory terminals, either on the apical dendrites, not drawn here, or on the basal dendrites. We have inhibitory terminals. So the excitatory terminals would be secreting glutamate. And the inhibitory terminals would be secreting GABA, or in some cases glycine. And then we have the axon, including the myelin in many CNS neurons, and the space at roughly one millimeter spacing is where there's nothing. So it's called a node. And the question is, who is the scientist who discovered the node of Ronvie? But I'm learning my code. Okay, so then the axon bifurcates and synapses onto a postsynaptic cell, where that's now presynaptic terminals, and we have the whole process that goes on again. Any questions? So as usual, we like to take a nucleus or a region of the brain and emphasize that region of the brain. So we've discussed the midbrain in the context of nicotinic receptors, and we've discussed the hippocampus in the context of glutamate receptors. Today we're going to talk about the cerebellum in the context of GABA-ergic transmission. There certainly is GABA-ergic transmission in the midbrain, the dopamine-ergic neurons, and there certainly is GABA-ergic transmission in the hippocampus. But in a sense, the most regular nucleus in the brain, the most regular structure in the brain, is the cerebellum. Here it is in most rodents. It's in back of the cortex. Here in the human brain, it's actually below the cortex, and here's the spinal cord. It has highly regular structures, highly regular layers, and a stereotyped set of projections from one cell to another all over the cerebellar cortex. In fact, one of the cell types in the cerebellum, the so-called granule cell, contains a plurality. It represents a plurality of the cells in the human central nervous system or in any central nervous system. Not a majority, but it is a more common cell type than any other cell types. In fact, probably 10% of the neurons in the central nervous system are cerebellar granule cells. Well, if you have that many, they're all individually very small, which is why they're called granules. Those granule cells have dendrites locally, and the granule cells are located fittingly enough in the granule cell layer. Then there is another hero of the story, very few of these, but very large cells, the Purkinje cells, and you know who discovered the Purkinje cells. And then the outer layer of the cerebral cortex is called the molecular layer, not because it keeps molecular biologists busy or that because that's where molecules live or that's because molecular biologists live. But in the old days, with just light micrographs, there were all of these little dots. People said, oh, this is the smallest component, so we'll call it molecular too bad, but that's what it's called. And then down below, projecting to other regions of the central nervous system is white matter that is myelinated axons. Typically, white matter in the brain, not the one that you saw that Ralph passed around that was pretty well fixed, and you couldn't tell. But in most cases for unstained brains, myelin is much lighter in color than the cell bodies. So there's this wonderful circuit that has really played a large role in understanding neuroscience. The Purkinje cells, those giant cells, are quite interesting because you can think of them as highly elaborate hands. If you look at them face on, the dendrites ramify over hundreds of microns, all within a given plane. Those dendrites all intersect the axons of the granule cells up here in the molecular layer of the cerebellum. But if you look at them sideways, all of those hands line up this way and they look very thin.  The cerebellar folia do twist and turn, so what is face on from one view may be sideways from another view, but this structure on a local scale exists. Here are in an electron micrograph is the molecular layer. Each of these slightly sub micron diameter structures, roughly half a micron, are in fact the axons of individual granule cells. These are not synaptic vesicles, they're axons, unmyelinated obviously, packed as closely together as they can be. Any questions about the structure of the cerebellum? Obviously highly interesting and entertaining structure. But as Ralph Adolf said, as interesting as the cerebellum is and is regularly organized and as well studied, a somewhat embarrassing point is that we're not actually sure what it does. So people have lived without the cerebellum. And animals live without the cerebellum. Obviously it's there to fine tune one or more functions of the nervous system, probably sensory, but we're not sure exactly what. Now, candle shows in one of the early illustrations in chapter 10. He shows types of synapses. There's type one on a spine. There's type one on a shats dendritic shaft. And then there's a type two on the cell body. So type one is. Axo dendritic. Sorry, type or Axo spine. The type two is typically exosomatic. Now, if you look very carefully in the electron micrograph, you can see differences between the type one and type two. The synaptic vesicles in the type two are oval and in the type one they're round. That's pretty much an outmoded terminology. So we're not going to mind the type one and the type two stuff. But we should know that there are synapses on dendrites called Axo dendritic. And synapses on the cell body on the soma called exosomatic. Okay, so we're not going to emphasize the type one. All right, so we have a general view of the synapses in the nervous system, but they look like where they are. Now we're going to talk about integration of signals by synapses. Remember the typical CNS neuron has about a thousand synapses. In most cases, the output of a synap of a neuron is a firing pattern. And so the neuron really integrates all of those inputs and either does fire a spike or a train of spikes or does not. And to some extent, this depends on whether those inputs exceed threshold or don't exceed threshold for firing a spike. Everyone clear on this? Okay, so there are several types of synaptic integration. There is temporal integration and temporal integration between synaptic inputs itself can have two causes. It can be caused by molecular lifetimes and it can be caused by the fact that the membrane is a capacitor and filters signals spreads the mountain time. We'll give examples of that. Then we can have spatial integration. Think of the Purkinje cell with all of those dendrites. So we would have integration between an input on one of the dendrites and an input on another dendrite and the cable properties of those two dendrites would play a role in how they sum. And finally, we can have what we have learned today. We can have integration of excitatory and inhibitory inputs so that in the absence of inhibitory inputs, many excitations would get to be super threshold. But in fact, because of inhibitory inputs, many fewer get to be super threshold. And there are regions of the brain where a majority of the synapses are in fact inhibitory. Because there are so many excitatory granule cells in the cerebellum that's not true in the cerebellum, it is true in regions of the midbrain. Now, so let's go over the method, the kinds of integration. In a previous lecture, we talked about synaptic integration due to molecular lifetimes. That is, the concentration of acetylcholine at the nerve muscle synapse is a delta function in time. It's less brief than a millisecond. And yet the current in the synaptic cleft or the current due to the receptors is longer than a millisecond because those channels all get opened nearly synchronously, but they close stochastically with a time constant of about a millisecond. We talked about the transition from an open state to a closed state, and we said that that transition is rather leisurely, at least on a time scale of milliseconds. In some cases, channels stay open with transmitter bound for a millisecond, others for 10 milliseconds, and MDA receptors even longer for tens of milliseconds. That all has to do with the transitions between open and closed states and the eventual dissociation of the transmitter. All of this works like a single exponential because of acetylcholine esterase, the enzyme which hydrolyzes acetylcholine into acetate plus choline and is very, very efficient. So as molecules of acetylcholine leave the receptors, acetylcholine esterase can take care of them and hydrolyze them. They don't act again as long as acetylcholine esterase is present. If acetylcholine esterase is not present or it has been inactivated as in some insecticides or nerve gases, then the consequences are very severe. Okay, so we've talked about integration due to molecular lifetimes. We've talked about the units. Now, that was the story for acetylcholine esterase, which is present at high enough densities so that it can hydrolyze the acetylcholine. There is also usually a delta function of glutamate and GABA at CNS synapses. Now, there is no enzyme like acetylcholine esterase at CNS synapses. Nothing hydrolyzes GABA and glutamate. Instead, the GABA and glutamate signals are terminated just about as rapidly as acetylcholine by transporters. In fact, by sodium coupled transporters, which harness the difference between extracellular and intracellular sodium to drive GABA or glutamate. And those two are present at densities of more than a thousand per square micron. I'll show you how we know that later in the talk today. Here in this schematized view of the CNS synapse, we have the glutamate transporters. Actually, not surprisingly, there are several genes for glutamate transporters. Some of them are actually on presynaptic cells. Some of them are on glia. The same is true for GABA transporters.  Those transporters that are on presynaptic cells obviously help to replenish the transmitter, which then gets pumped back up into vesicles. But there's also this very efficient elimination of transmitters that occurs on glia. It can't be reused, but it does sharpen up signal transmission in the central nervous system. So we have these delta functions of transmitter, which typically produce inwards synaptic currents. And then we have the capacitive filtering that simply is due to the fact that once you put charge on a membrane, because the currents have flowed and charged up the membrane, it takes a while for that charge to leak back off the membrane, and therefore the voltage for the depolarization to decay back to zero. So the simple fact that membranes are RC filters and have time constants spreads out the synaptic potentials and allows them to sum more distantly in time than one might think naively. So here we have temporal summation in a cell. We have two synaptic currents, each of which are relatively brief about a millisecond with the decay that I told you about. But the synaptic potentials can have time constants in some cells as long as 100 milliseconds. So even though the synaptic current channels remain open for about a millisecond, the voltage in the membrane stays depolarized for tens of milliseconds. So if we have two synaptic currents, they can add partially so that the second one can reach threshold, even though the first one, by adding to what remains from the first one, even though they are so separated in time. If the membrane time constant is briefer only 20 milliseconds, for instance, then these two synaptic potentials would not sum with each other. So the concept of spatial summation in a cell for good or for bad depends strongly on the passive characteristics of the membrane. Any questions about this? And then we can have spatial summation. So here we have a cell body and it's usually easier to place an electrode in a cell body and to record from that cell body than it is in the dendrites, although later on in the talk I'll show you that you can do that. And also important from the encoding properties of a neuron. So if we have a synaptic current at point A and a synaptic current at point B, there is a possibility that they can sum right here in the soma if the length constant of the cable is long enough. Typically in a neuron, the length constant is on the order of a millimeter, sometimes half or a third of a millimeter, usually not very much larger than a millimeter. And so that will determine whether or not those two inputs on the dendrite can sum to reach threshold. So a short length constant does not allow them to sum a long length constant does allow them to sum. We're doing okay so far. Now actually there are additional delightful complications. If dendrites were completely passive, they would act like leaky cables. So if for instance, but as you'll see they're not, but let's assume for this slide that they are, if we put an electrode in the soma and have an excitatory postsynaptic potential right here near the soma, we get an EPSP, we can measure it, so that measured in the soma, the EPSP decays and is much smaller out here. And so we have a length constant of something like a millimeter here. On the other hand, if we have an input here at number six and measure the EPSP, then locally the EPSP is much larger, but we won't be able to measure it very well in the soma. So we get this peak right here and then it decays pretty rapidly. Any questions? Okay, so with these concepts you would expect dendrites to passively integrate inputs without any fancy sodium channels. So if we have two simultaneous co-localized excitatory postsynaptic potentials, then we expect them both to look like this in individual trials. One is a little larger than the other. On the other hand, if they sum, if they occur during the same trial, they can sum pretty well and they might get over-threshold when recorded in the cell body. If they occur simultaneously but at different places on the axon, they can also sum in the cell body, although the rising phase is a little more broad, and they could, so these are spatially distinct. And so it's possible just using the fundamental equations of electricity to simulate stuff like this. And there are lots of programs out there in the literature that do that. So here we have a cell body with three dendrites. This is a simulation program called Neuron, but there are others. And what we do is to have these three simulated inputs, they are occurring at different times, and one of them is able to produce an impulse. Let's run it again. Check on your browser, click on your browser's refresh button. Let me decrease the, let's see if clicking on refresh works. Yes, it doesn't. We have to go back, and this is the current, the voltage in time, and this is the voltage in space at various regions along the dendrite and parametrically in time. So let's run it again. Big depolarization was an action potential. So the more we know about the characteristics of a neuron and the more we know about the ion channels in that neuron, the more we can simulate about the encoding properties of that neuron. And there was just an article by a group in Europe called the Blue Brain Project, which is simulating an entire column in sensory cortex, all of the neurons in that column. But the fundamental units of that simulation are very much like the ones that we have talked about today. There is a complication and this complication is that the dendrites are not passive structures, simply with resistors and capacitors and with neurotransmitter gated channels. Actually, if you look at them more carefully, and you can do that now because you can fill a cell with a fluorescent dye by patching onto it with a, onto the soma with a patch pipette that contains a fluorescent dye. So here's the soma. Or you can light up all of the branches of the cell by having just that one cell or just a few cells express GFP, and so everything in the cell will light up. And then, of course, this is done deep inside a several hundred micron diameter slice of brain. These days it can also be done in a living animal, but the success rate is very, very low. But if it's done correctly using two photon microscopes, you can image the dendrites of a neuron. Not yet it's axon. The axon is really too thin to be visualized correctly. But you can visualize the primary dendrites of a neuron and therefore come in with a patch pipette and come in and patch on the primary dendrites of a neuron a few microns in diameter and therefore record voltage from those primary dendrites. And lo and behold, there are action potentials in those dendrites and in some cases they overshoot from over zero. And so it's pretty sure that there are actually electrically gated sodium channels in those dendrites. And to be absolutely sure, here's an experiment in which a person patched onto a dendrite, did not break into the dendrite, but kept the patch pipette extracellularly and recorded single channels. And so here now you're only depolarizing a local region of the dendrite, not the entire dendrite. And it looks for all of the world as though when you depolarize in the, and I showed you simulations of this sort of experiment a couple of weeks ago, and when the experiment is done with depolarizations, you do see individual single channels. These are blocked for instance in the presence of tetrodotoxin. And when the experiment is done repeatedly over a time course of several minutes, the waveform looks just like the waveforms of voltage clamped sodium channels. It depends on the depolarizing voltage. Here is a small depolarizing voltage. Here's a large depolarizing voltage. Here's an intermediate one. So clearly there are sodium channels in dendrites. They are certainly at the axon hillock. In fact, the region of the axon hillock, which is going to give rise to the axon, is the densest sodium channel population. But out here on the dendrites there are also sodium channels. And so that means that voltage gated sodium and calcium channels in dendrites allow the action potential to back propagate into the dendrites and produce much larger signals than they might otherwise. So here is an overshooting action potential at greater detail in a dendrite. You can't patch individual spines, but you can patch the dendritic shaft. And so this implies that parts of cells can actually process signals semi-independently. So not only does the neuron itself make a decision about how to fire a spike and go to the next cell, but the neuron can also have individual isolated parts that process signals. So this is very exciting, and we should stay tuned to see how this works out. Now, let's talk about the veto principle of inhibitory transmission. Because of the way currents flow, inhibitory synapses work best when they're near-electrically the excitatory event that they're going to inhibit. Near means less than one cable length. So this means that on dendrites inhibitory synapses do an excellent job of inhibiting EPSPs, and excitatory post-synaptic potentials on nearby spines, but typically not on far away spines, perhaps on different dendrites. And it also means that inhibitory synapses on cell bodies and initial segments do a very good job of inhibiting spikes wherever they might arise in a cell. So in some cases, there's a selective advantage of local excitation and inhibition in a part of a neuron. In other cases, there's a selective advantage of vetoing everything in a neuron by an inhibitory synapse. The best example of a veto, well, one example of a veto, we're going to move away from the cerebellum for a couple of minutes, but in the cerebral cortex, there is the so-called chandelier cell. The chandelier cell is an inhibitory neuron. It makes GABA. You can see why it's called a chandelier cell. It seems to have these little candles. So this is one cell, one chandelier cell. It's axon branches. The axon is this little thin thing here. And it branches many times, produces these little candles at many different pyramidal excitatory neurons. So here, this is the schematic of this. You can clearly see the dendrites of the chandelier cell. There are apical dendrites and basal dendrites. But the axon is this very thin branched device here. And the axon makes these cartridges on the axon hillock, lots of pyramidal cells. The axon hillock is where spikes arrive. So this is a great example of the veto principle in which an inhibitory neuron can prevent an excitatory neuron from firing spikes. And in fact, this inhibitory chandelier cell can prevent many excitatory neurons. Simultaneously from firing spikes. Any questions about this figure? So there are veto synapses in the cerebellum, back to the cerebellum. The most obvious veto synapse would be right here on the cell body of a Purkinje cell. And that is where we're going to look. In fact, the inhibitory veto GABA-ergic synapses are so obvious around a Purkinje cell. Here, you can imagine that there's a Purkinje cell surrounded by these terminals. Actually, in the light microscopes, they look like paintbrushes surrounding the cell. And so the French neuroanatomists who discovered this called the pesso or paintbrushes. So we are going to study, we're going to show you some diagrams now. of GABA-ergic paintbrushes or pesso. To do this, we're also going to address the question, the conclusion that I gave you about 20 minutes ago, which is that there are large numbers of GABA transporters, large enough, locally dense enough to remove all of the GABA when it has, after it has acted on a GABA receptor. So here is a fusion protein between a GABA transporter. At the time these, actually it's still true that nobody knows the three dimension, the X-ray structure of the GABA transporter, but we know it's topology well enough to know that there are 12 transmembrane domains, which way they go, which ones are important. This particular GABA transporter has been fused to a molecule of GFP, even though it's blue here, take my word for it, it's a molecule of GFP. And this allows one to visualize the GABA transporter wherever it occurs. And this is a knock-in mouse, a transgenic mouse, whose GABA transporters have all been replaced by GABA transporters fused to GFP in the head. So here is a sagittal section, remember the sagittal plane is this way, it's of the head of the mouse, the central nervous system, the brain. Here we have the olfactory lobe. Here we have the cortex, which in a rodent is not too impressive. Here is the hippocampus looking like the curled seahorse. Here's the midbrain. Here's the cerebellum, the hero of today's talk. Here's the hindbrain leading to the spinal cord. So you've heard about all of these structures. They're all fluorescent green in this knock-in mouse containing fluorescently labeled GABA transporters because there are GABA urgic synapses and GABA transporters many different places in the brain. Any questions about what this slide means? So what we're going to do is to zoom in on the cerebellum. That's where we want to be counting the density of GABA transporters. So here is a individual folium of the cerebellum. And you can see, if you concentrate on this cerebellum, that there are layers of bright green. Those are really the cellular layers and layers of background. And that's the white matter. That's where the axons are, not the GABA transporters. If we zoom in a little more, here we have the... This is an antibody to the GABA transporter. Not very quantitative, but nonetheless it gives us an image of where the GABA transporters are. Here in the granule cell layer, below the purkinje cell layer, there definitely are GABA transporters. You can see some reticulate gray here, not so many. Here is a purkinje cell. There are five of them in this picture. Here are the pesto, the paintbrushes, which are inhibitory terminals containing GABA transporters to take that GABA back up. It stains very heavily. Here's the molecular layer, the so-called basket cells, which are also our GABA retic. And so what we've done here is simply to take a negative photoshop the image, so that the parts that stained most brightly are now lightest. And here is a light micrograph of one of the knock-in mice with fluorescent GABA transporters. The great advantage of having the fluorescent GABA transporters is that they can be counted and quantified much more precisely than immunocytic chemistry. But you can see that the general localization seems to agree well between the antibody staining and the GABA transporter. There is a little bit of reticulate staining in the molecular layer, lots in the pesto, and a fair amount in the molecular... sorry, a little bit in the granule cell layer, lots in the pesto, and a fair amount also in the molecular layer, which is because there are lots of inhibitory terminals on the dendrites of the Purkinje cell. And in fact, if we zoom in and look at the dendrites of a Purkinje cell, we're zooming in again, what we see is the presynaptic terminals of GABA-ergic neurons. Now you can see why synaptic terminals are often called butons, because they're like little buttons. So this now is 50 microns across, and the butons are on the order of a micron or two across. These light areas that look a little bit like the Milky Way are actually the radial glia cells, which Ralph told you about, that serve both as functions of glia and also developing neurons migrate along them. You can see that the butons are connected in many places by strings of green, so actually each presynaptic basket cell has an axon, and on that axon are strings of butons. The axon itself also has GABA transporters in it. And so it is now possible to use quantitative confocal microscopy, calibrating this with individual fluorescent GFP molecules to show that the transporter density is around 1,000 per square micron. And if we now zoom in on an individual buton, well, that's interesting too, but not in the context of today's talk. We'll stop here. So my office hour is as usual. The red door. See you on Wednesday.


So I came across this company called Kicks Case and I saw they had a bunch of cool designs when it comes to the bags and the colors and prints, but I wanted to see if the bag was actually even worth it. So in today's video, we're going to be putting my size 13 shoes inside this bag, talking about the functionalities and all the things that come along with it, and I bought a couple extra bags to not only show you guys the different colors, but to give one away to you. So make sure you stick around to the end of the video to find out how to win one for free, and if you didn't know by now, my name is DJ, and this is the DNA show. On this channel, I love talking about sneakers, giving you guys in-depth reviews, and especially helping you guys know the ins and outs of the different accessories that come along the way with being a sneakerhead. So if you haven't already, consider subscribing and joining the fam. We're on the road to a million subscribers, and you could be the next one to get us there. So starting off with the bag right here, as you can see, you have a zipper on the front end, and you can stick your hand in here, and this goes all the way down to the bottom of the pocket. So this is going to be a convenient area to put slim things that you may have as it's going along in your travel bag. And then on the side right here on both sides, you have a zipper that comes up, and then two bigger compartments right here that fold out. And these are actually, I would say, about a couple inches wide, and then obviously the whole height of the bag. So this is going to be a cool compartment. One good thing that I like about having these side pockets on the bag is, yeah, you can't put a pair of shoes in there, but you could put, you know, extra clothes or towels or smaller treak-ins or little items or something like that, because this is typically a bag that you're going to take for carry-on. Based off of the dimensions of this bag, it is TSA carry-on approved, which means you can put it in an overhead bin, and then you can save money and not have to worry about paying for luggage, which is always another bonus as well. Now looking at the zipper right here, you got two zippers that come around, and then you got the flap that comes over, and then you got the shoes inside. Oh, you got the shoes. So as you can see right here, I have four different pairs of sneakers, and I am a size 13, so I have some Air Max 90, some Air Jordan 6s, some Kobe Zoom 1s, and some Nike SB Dunk Lows. So with that being said, typically an Air Jordan 6 is a very bulky sneaker, and this is a shoe that I can fit in very easily and not really have any problems with that, so that's one thing that I really like about this bag, because when you're a size 13, it's definitely a struggle, fitting your shoes in suitcases and backpacks and different luggage and stuff like that, so for me, I always want to make sure that I'm carrying all my sneakers. That way, I can make sure that they're protected along my traveling process, because you could easily have your shoes get crumpled or something like that when they're in your suitcase or getting tossed around, and in that way, you can make sure that these valuable sneakers are always taken care of. So as you can see right here, you have three different dividers as well. These things are removable and adjustable, so basically they're just Velcro strapped on the inside, and then from there, you can move them around. Maybe you put two pairs of shoes in something here or three pairs of shoes or one pair of shoes and then a big open space.  Another cool element to this bag in, I'm probably sure you didn't see it already, but there's actually a whole laptop slot right here in the back end, and this just slides right in behind the sneakers on the inside of the bag. So that's cool as well. Typically, I'll take two pairs of shoes, like I have a pair on my feet, an additional pair, or sometimes I'll have that third pair, depending on where the trip is and where I'm going and what I'm going to be doing and all those different factors. But that usually leaves me enough open space to then put some camera gear or some clothes. I always like to put my hat to my carry-on bag. That way, I never have to worry about them getting crumpled up, and every single time they're always protected. So that's another good thing for these type of bags as well. Now, looking on the lid of the bag right here, you have three different zippers. So there's going to be one that goes behind this compartment, whole area right here. Then you have a smaller one right here, where you can now stick something small in here, and then you have a bigger one with the net, with the mesh where you can see everything here. Typically, I use these compartments for what I'm adding, like my phone chargers, sneaker wipes, batteries, different things for my camera gear, my hard drive, stuff like that. So it definitely makes it a lot more convenient when I can put my little items inside of here, and they're not rattling around all over the bag, or they're easily accessible. So that's always a cool touch as well when I see bags like this. Now, when it comes to the handles and things like that, I'm going to switch up the bag for you guys, just to show you guys a couple of different color options. So this one's actually kind of cool because it has a split, and you got that red camo with the black camo, and then it's got the case case right here. And it's also cool because if you look really close on the zippers as well, it says, case case as well. So there's a lot of nice little touches and details. You know, typically when we're looking at sneakers, right, there's always cool details to each shoe. So when it comes to this bag, now there's gonna be different color options on the outside, but essentially this is gonna be all configured the same when it comes to the compartments, the zippers, and everything like that. So typically, if you look at the website, which I noticed, most of them are essentially all the same bags, just different colors based off of your preference, which is a cool thing because some people might like that olive. Some people might like this crazy camo print, or they might want something more simple with just the black bag.  So I think when it comes to quality, presentation, creativity, and overall functionality, I feel like they're doing a solid job when it comes to a bag like this, especially to say that these are easily under $100, and that's another cool thing, right? Because typically we see a lot of bags that are $100, $150, $120, and these right here you can find for around 80 bucks, and if you use the discount code DNASHOW, I made sure that I got you guys 10% off as well, so that's another perk for you guys if you wanted to save and maybe get the bag for around 70 bucks or something like that. So I think when it comes to bang for the buck in style and everything like that and having a cool travel bag, this is definitely gonna be a really good option. Another thing that's really cool that they have that came with one of my bags is this has an additional bag that goes inside of there that fits inside the slats, and you can unzip this whole thing, and now you can put something like in here, maybe dirty socks and drawers or toiletries or whatever you need to do to make sure it's not rattling around, a bunch of kind of smaller items, different things like that, so it's cool to have these little bags as well, especially if you just wanna carry this around, walking with you on a trip or something, you need to go somewhere or you're going to an event or something like that. So to have this in addition with one of the bags or to buy it separately and just have one of these, these are always convenient as well, even if you're throwing it in your actual suitcase. Also with these, you can fit a smaller size pair of shoes in here, now this is a size nine pair of shoes, I'm a size 13, so it's not really gonna fit unless it's like a really slim sneaker, but based on the sizes, it's gonna be really, really tight, but I think if you have a size, obviously anything besides probably a 13, maybe a 12 and under, I think you shouldn't have too big of a problem putting a shoe in here unless it's like an extremely high top shoe or Jordan one or something like that, so that's something always to consider as well, if you have more of a premium type sneaker that you really don't wanna get messed up and you wanna keep it protected in a bag like this, definitely very convenient. So I'm sure you guys are wondering at this point, how can you win one of these for free? Now again, like I said, if you place any orders on their website, you can use the discount code DNASHOW and that'll get you 10% off of your orders, and if you wanna have a chance to win one of the bags, you're gonna have to be a part of the DNAFAM, so I'll have the link for you guys down below as well, and basically all you're gonna have to do is accept the challenge, that will submit your entry, from that point you'll be good to go, and then I'll select the winner at the end of the month, which is gonna be in a couple weeks, and again, the discount will be always valid for everybody who wants to use it, because I know everybody can't win one for free, but at the same time I wanna make sure you guys can get at least some type of discount. So I hope this video was helpful, and if you ever wanna see any other types of bags, accessories, cleaners, whatever it may be, related to being a sneakerhead, let me know and I'll try to get a review for you guys, that way you can see the whole product before you actually decide to purchase it, because I know a lot of people, they're always like, well I don't know if I should get it, I need to see something a little bit more in depth, so hopefully I did a good job showing you guys what it looks like. I really like these bags, especially the fact that they got all these cool colorways, and when I saw this one, I was like, bruh, low key, this is about to be my new travel bag, so I'm definitely taking this on my next trip. Yo, if you enjoyed this video and wanna grow your collection, or make extra money on the side, I built a VIP mastermind that will teach you everything that I've learned about growing my sneaker collection over the past 15 years. This will also give you access to the DNA fam and my VIP community, where we talk about investing outside of sneakers. And don't worry, if you don't plan on joining the VIP community, it's okay. I also set up a private DNA fam community that gives you access to all the behind the scenes looks from the studio, and multiple chances to win free sneakers and gear from weekly and monthly challenges, so all you need to do is click on the link down below in the description, or the first link pinned in the comment section. That will get you set up and into the community, I'm excited to see you guys on the inside. If you made it to the end of this video, let me know one of your favorite travel sneakers you gotta always have with you. For me, it's a presto. I don't care what colorway, but I gotta take at least one pair of prestos with me on a trip.


should not have worn red. Hello everybody oh Ty you should join in on the live so that if we get questions on our live we can read them because sometimes this thing goes so crazy. What's up guys so today yeah are you speaking? I'm at my house and I know it's been a while since you guys got to chill with me at my house and with the birdies so I wanted to give you an update on how Josie is doing hey hey Caleb and Leo and uh like who who Josie has bonded with and all of that hey Divya it's been a while hey Apollo so um I'm I'm totally happy to what's up peekaboo cockatoo I'm totally happy to talk to you guys do Q&A whatever you guys want to do I figure I'll be on here about like 20 minutes or so um and you can thank Ty because Ty forced me to go Ty literally was like I'm coming to your house I'm getting the birds ready and we are going to go live because Ty is uh has been a viewer of the channel so hey Tracy hey Lisa so he he's kind of like has the sensibility of you guys Jersey we need to bring Jersey so Jersey does this thing for those of you don't know who Jersey is she is a cockatoo hi Isaac and she can get uh I don't want to say moody because she's always really sweet but sometimes she doesn't want to like stand like that is she doesn't want to come to me and or anybody and then she'll let you kiss her I'll be like can I have a kiss and then she'll be like she'll kiss you she's very like polite but then she'll like not step up so today when I try to do it if you guys remember and saw the video of Josie she is an amazon parrot she's a yellow-naked amazon and she is she could be about 45 years old they said when they did the math uh up to about 45 definitely no less than 35 and um I don't know if you guys remember I said she loves guys well the moment she she's in love with my brother but the moment she met Ty she fell in love with Ty Ty's already like did you say you already took her to the mall yeah part of the mall yeah Ty likes to run around with animals at the mall and especially his uh his chameleon lifesaver who's like the most gorgeous chameleon you'll ever see in your life but um so she's doing well now the last time I did a live and talk to you about Josie I told you that she liked men more and at that time because things keep changing you always like learn about a bird when you have a new bird it takes like there are animals like you know dogs can be a little easier to read right away birds can keep coming around and around like sometimes for years and who you thought the bird was can be completely different in two months than who you know the bird to be now you understand what I meant but anyway so Josie was very much all about guys and she also I thought she pretty much hated me honestly because she knows what you're talking about yeah she's just looking because when I would go to pick her up she'd be like and uh I kind of thought maybe it's because when the day that she came here she was you know I was the one that was kind of like the first one she saw so like maybe she was like she's the one that stole me if that makes any sense hey love bird love oh yeah and also I saw Megan and Divya um so but now uh we were discussing this like about an hour ago she is very much what she does is she plays hard to get so for example I'm not going to be her favorite person and you know how birds have tears right so if I'm the only one around she might come to me but then uh but then like if a guy is around um second tier it's not happening um so when I am around now I found out that if I try to take her out of her cage I mean her cage is always open she doesn't appreciate it so much but if I go up to her and I call her fancy and I say hi fancy and then I um just kind of leave her alone she'll climb up to the door and then she'll tell me two minutes later hi bird and I'm like okay she's like ready for interaction if that makes sense um Tracy G it's it you have a good eye Leo is stressed um there's been some changes you know we've been building a birdroom George called me today and said hey I just want to you to know that I want you to like take a look at Leo I think like he's he's stressed he might have alluded to he thought he was sick and I was like panicking and I wanted to come home right away but I was waiting for the mailman to come with the boxes and he did not come the mailman and this is the stuff I have to deal with but I um I got home and he's he's actually fine he's just he's just stressed he's not sick or anything like that I think he's stressed because while we're you know working on the bird room we're not actively working on it while they're in there but it's an adjustment while we get them everything that they need in there just continuing to improve upon it and also I think they're very used to being like in the kitchen and everything like that so right by us all day um oh wow thank you for the oh let me see it how do I was here oh okay oh Ty's got it ready for me captain third leg thank you so much for the family what do I say thank you so much for the what do you call this oh no my words are not been here recently the the chat what do you guys call it Megan I know you're online I forgot what the super chat the super chat okay I have a cute cute clingy parrot lip but she likes to nibble my ear how do I make her get over my ears or my lips oh guys this is a hard question I'm gonna tell you this right now I think about this all the time it really is a great question yeah Leo Leo saying because I have Nelly my Senegal she does this to me she's always biting my face and you know with birds with other behaviors where's the trap master yeah the trap master you know with birds where it's very much that you um it can you screenshot this with your phone for a second uh you know with birds where it's very much that you got to make the adjustment not the bird and you can kind of redirect and and you know use positive reinforcement with this situation there are a few things that I do but I don't think they actually solve the problem and getting the bird to stop doing that now one thing you can always do though is recognize why the bird is doing it and start to become in tune with your bird and be like okay if I don't you know scratch her head every five minutes she's gonna peck my face and that's pretty much what it is with uh Nelly right is that Leo it is Leo so what happens with Leo no Leo whoa my words like guys I've had brain fog now so uh so what happens with Nelly is she'll come and she'll peck at my face it's not so bad but it's really annoying right so after analyzing her a little bit I started realizing that what she is doing is trying to get attention so while she's on me I'll sense like okay every couple of you know every two minutes or every five minutes let me just scratch her head before she does that now of course the thing with that is I'm adjusting to her that doesn't necessarily teach her not to do it but what it does do is get her used to not doing it so for example if if I'm doing that all the time and she doesn't have to tap my face for that kind of attention or to feel validated then she'll forget that she does that um but it's a consistent thing the other thing like hacks right oh I'm sorry um I literally my vinegar gang hoodie and our new hoodie like okay that's why I love hoodies okay like I like wear my hoodie and then put it up and then it covers like this bit of my face you know and those are just like lazy hacks like if you if you're lazy and you're like you're like I just I just like I don't I don't have the arms space to like you know but the main takeaway from this is what you want to do is figure out the reason why the bird is doing it and see if you can prevent the bird from doing it by making the bird feel more comfortable about what it is that they need does that make sense um I hope that makes sense you can ask me more about it if there's something else you want to know um but that's a great question I I think about that all the time because it's so annoying like I know I I totally feel you oh my god guys doesn't Leo's red I don't know if it looks like this to you but it is like it shines George has some strange color lights I think and and they like he goes like purplish red um what does that say okay bonding with a bird is not really something that can be answered in two minutes it's really a long term commitment in terms of your commitment to bonding so for example one of the first things I do when I get a bird that you know may not like me or may not like anyone is I include them in everything they do in the best way that they feel comfortable so what that means is let's take Josie for example I could get her to step up and um let's say she doesn't want to step up like I get to the point where I'm like no you you are going to step up and me and you are going to the shower and then when I go work on my computer me and you are going and take a different bird like let's say Rocky I can't have him on my shoulder he I'm not comfortable with him like that he's not comfortable with me like that so I will take him let's say to the shower but he'll be somewhere else on a perch or I'll roll his stand into the living room with me until they it's called habituation so they get like habituated to be around you and that is that is like it becomes their norm then they're just getting adjusted to it honestly it's like it's like munchausen no not munchausen what's the one where you fall in love with your captor uh Stockholm Stockholm Stockholm syndrome right like you're providing everything that they need um I just I know that sounds crazy but I've thought deeply about it I mean these birds are not meant to be in our home right but you gotta make them feel safe you gotta make them feel loved and included in the best way they feel comfortable another example is my Indian ring neck blue blue is borderline wild in the south okay blue does not want to be picked up and loved and kissed and all of that and guess what that's okay like I'm not here to force him to be like that I kind of like it I like that he feels wild and that he has brando to be wild with and be a bird um however he knows the rules like when I tell him to go to bed he doesn't want me picking him up and touching him he'll fly to bed um he'll just go to bed because he's like hell no I don't want her like trying to get me and catch me so he's like I'm just gonna do what you told me to do and um it's it's it's it's amazing it's amazing to see a wild like a bird live kind of wildish you know but he will he also what's fascinating guys is like blue's not the kind of bird that I can really put my hands in the cage and he's going to step up and be like I'm so excited to see you and like he wants to kiss me and all of that and but let's say I want to put him outside in the aviary and I want to pick him up and brando is going in there I can put my hand in the cage and he will step up to me he will step up and then I'll just grab him and like carry him like this and he he he knows like he knows where he's going and why he's doing it so I'm adjusting to him so as long as he has his freedom and I'm not trying to force him to be the bird that I want him to be I'm letting him be the bird that he is and that that comes a long way now if I didn't have brando and he wasn't in love with brando and he didn't have a fellow you know mustache parakeet ring neck like to hang out with he would behave completely differently than I would be what he relies on but I love birds being birds they need to forage they need to chew they need to nest you know like I don't believe in not encouraging birds to be birds I don't believe in taking all these things away from birds so that we can live with them like we have to try our best to provide you know like what they they would you know have in the wild you know huh they need to steal your stylus my stylus is right here guys brando knows how to pull it out of my phone okay and then fly away with it I lost a few of those already what else you guys got what else you guys do you can you see like I don't know what it looks like but on my phone I'm in the light you could see it looks like Leo's glowing okay I knew that was going to happen she is on her favorite person if you approach a bird on their favorites okay he's he's getting stressed because I paid her attention so that's something you got to know about your birds you know um yeah he does have super bright eyes um so if you guys noticed that like she wanted to you know bite me or warn me but it's because she's on her favorite person and I just gotta know you don't approach a bird when they're on their tier one person that's it and um look look how happy she is and I just don't you know but let's say you have a bird in the house that is very much like this you got a bird it's your dream bird and you bring it home and it falls in love with your significant other happens it happens all the time and the issue is that most people give up they're like ah he loves you more so I'm not even going to put the effort in and now you have to put more effort in and more work in to work with the bird and what you have to do is you have to work with the bird when the significant other or that person is not in the house at all like at all because they are so loyal and they will not want to act like you know they're cheating on their their favorite person um so that's the first thing you got to do and then you got to put a lot of effort into including that bird and everything you do alone if the bird is food motivated you feed them you know you give them all the like treats and all the excitement especially amazons they're very food motivated how do I get my bird to what come to my hand I don't know why this stuff disappears so come to my hand he loves my shoulder but hates my hand and flies straight from my hand to my shoulder well at least you have a bird that loves being with you so that's the first thing the second thing is uh I don't know exactly what kind of bird you have and a lot of times like that actually matters they each species has different like tendencies that it's like oh it's an amazon I know what to do or oh it's a ring neck I know what to do things like that so did you answer that I feel like um no answer there but I think I saw it I'm not sure so um conure it's a conure okay so have you worked on stepping your bird practicing stepping up or does your bird bite you see because this is the thing guys it's much easier to do this kind of stuff like one on one because I have to kind of wait for the answer and know so much about the situation because every bird really is different so if you work with step up with your bird and you can get your bird to step up you can practice that with a tree you can practice that with positive reinforcement um and you you there's many different ways that you can do that with your bird but you have to get that bird practicing stepping up when you do that okay he likes to step up but after stepping up he flies to my hand so that can be in the motion and handling um kind of handling birds can be very much like sleight of hands honestly I swear and different birds just even different not just different species but different birds like you would require different things so for example let me see see look he's like hesitant he doesn't want to he's so happy right there okay baby it's okay it's okay it's okay okay um so for example sometimes like Leo let's say Leo was in front of me and I wanted him to step up and he refuses and he's kind of doing like the bite thing sometimes I'll tap their tail with amazon's this works pretty well because they don't move this fast and I'll tap the tail and they're like what is that let me step forward right and so those are just kind of like hacks that I use to make different birds move where you want them to go it could be a matter of literally you flattening out your hand right so sometimes let's say you're holding a like the bird is sitting like this and then you kind of flatten the hand it just gives leeway for them to go up to your shoulder so you really want to keep the bird steady um if you've been practicing step up and your bird flies to your shoulder that doesn't really sound like a bird that hates you but I don't know it's a hard situation to know over a live like this exactly what's going on I see that lovebird loft is actually messaging you and that's really good because I love that because then you guys are communicating and you might be able to notice some things that I'm not noticing here which I love and Annie uh I do I love our meetups at they just meeting you guys in person gives me so much energy and I love getting to know who you are and then later sending you guys feathered fun boxes or when you like order stuff like I remember you I'm like oh that's going to her and she has this bird and I met that bird and like um it's it's it's just really it really is fun like there is nothing like that anytime we've had a meetup I'm almost like what if nobody comes you know um sometimes we've been very last minute about them and you signed my bird no I wouldn't even people were asking me to sign the shirts yeah and I was like I don't want to mess up the shirt and they're like no it's okay I don't want to mess up our artwork but um can you sign the food dish the food dish oh we I I uh Megan's like mess up the shirt yeah she she knows Megan came to one of our meetups once right and you know what happens I don't know why all our meetups ended up being in mercury and retrograde because we know how I feel about that but communication yeah so I had ordered our rainbow bird shirts they are just gorgeous we have like the crop tops and a lot of times like I really do it from a local printer here I care how they look and I'll order different things too so that I can wear them and try them and if I don't find myself going back to it in the closet I'm not gonna wear I'm not gonna order it for you guys right so I ordered like one of multiple things and then the shirts that were gonna be at the meetup I think a few kinds but the girl that was supposed to help me be at the booth who worked for the printing company didn't show up so my brother had to jump in and Megan said hey do you guys have this shirt in and so my brother picks up another shirt and he's like this he doesn't know he doesn't know what a crop is and then she's like no like that but this and then he's like like this and then he had like the the wrong shirt and he's like let me just take your number and like ask my sister uh when is our next meetup Ty wanted me to have a meetup um yesterday when we did want to do it like this month right yeah we did but I don't know we should we have some cool stuff coming out next year oh my god guys hey when you say you're one of those that say next year and you mean like in three weeks in like 20 dude it's never tomorrow for me unless I've went to bed already like I like because I'm up past midnight every day so you know yeah that was funny you see that look she loves him brando so mad right now um I love Caitlin that you've been watching the videos what do you have what any tips on getting what kind of bird do you have to eat veggies so eat different birds I made the merch did you miss it we have it in pre-order right now I didn't promote it yet but I have it it's it's the the the landing zone shirt that you have been waiting for look at her nails her nails her nails match it yeah my nails match it did he's scared of her nails yes I've only told people that come to I didn't promote it yet we made videos but like I have to like actually edit them and put them together so uh we didn't like launch like let everyone know but we're gonna do a pre-order the reason we're doing uh pre-order is because it's four prints it's like expensive overhead and I don't know what sizes everybody's gonna be so that's what we're doing so everyone that gets in on this pre-order then they're gonna get their shirts fast because then again after that it won't if we do it again it won't be it'll also be another pre-order yeah Megan's been waiting for three years for these shirts and for these they match the nails I should have worn it it's like do I have it here I think Danny was wearing it Danny was wearing it I was like guys you don't want to eat though gorgeous you will be you will die go to parrotsation.com and look up landing zone type it in and see if you can find it literally we built our site ourselves and I don't know how to make it show up like it's not showing up in merch or anything like George was trying to help me and he's like you need to call customer service I'm like what so go to parrotsation.com and type in landing zone or even landing and don't finish the whole word when you see it come up if you see it click on it no Megan I am gonna promote it like I have we filmed videos but it's all up to me to like edit it the site does not show it in the merch exactly and I can't fix that that's why anyone that joins my live and sees this you guys are definitely getting the pre-order because like I haven't been able to fix that and promote it yet yes I was wearing it in a live you guys will you guys this is the most there is no bird merch like this none it is bad ass lovebird loft found it in a random way hey Wendy what's up um are you trying to do lives every week now I'm trying to and the reason being is I'm quite a perfectionist tie ties the reason ties the reason I'm quite a perfectionist and I want to get all video proofed hold on I'm going to answer that for you in one second para woman just one second on that because I have to answer that so I am going live because I'm such a perfectionist I want to have my videos edited and everything and I film all them but I don't really have an editor anymore so if anybody wants anybody wants to edit some videos so I'm I mean I can edit them more and ask but we're behind because we're trying to manage the site and do all this kind of stuff so then so then I'm like I might as well just go live because it's kind of edited you know like it's it's fun for me it that it's not doesn't do as well as regular videos where you could see the whole story but yeah um okay to answer that question what did you ask is Vinny upset about the you're not finding landing zone or zone landing shoot if anyone finds it can they put the link there is a link if you guys go to my last live I pinned the first comment and that is the link you found it is your corn still in the backseat yeah the corn is still in my purse you could just take the purse with you to the car if you want is it yes okay if any of you find the link can you post it for for for me because I I guys like imagine I'm trying to like like I can't even like pre-order it one of you asked me how is the how long is the pre-order so what we're doing is uh this shirt costs a lot just to make it's kind of insane so we are doing the pre-order so that we have like basically until we get a good minimum order and then we'll make them all and then I don't know because what we used to do with the Vinny gang is I'd have them already and then I'd sell them but then I'd sell out of somebody's size and not have it and then have to order a whole menagerie of them again um yeah we do have an Etsy store it's parrot station but the hoodie's not up there lovebird lops says if you type in hoodie you will see it I'm not sure if that's true um I could try on Ty's phone actually uh or Ty could try what you know if you bring that bird right here uh oh what's my favorite birch species I always used to say after the break Leo's upset uh but hey Nelly do you want to say hi to them look here I'll just have him sit on here what do you mean step up look at Leo he's like what did you do Ty what were you eating were you eating guys look at Nelly's eyes hold on oh Nelly's gonna show you her eyes um I wanted to answer what you said about Vinny does he like the new hoodie Vinny is pissed pissed um Vinny is no bird Vinny has never been scared of my nails I came on the first day and he was acting like Nelly looks huge she does look huge and they're the hoodie nails that's how big she is and I swear I swear he understands that it's a new hoodie and not his hoodie so I was like maybe it's not my nails and then the next day he was pissed again and he's been mad at me ever since and Ty said it's basically not my nails it's the design that's upsetting him Ty says it looks like movement or like it looks like it could be a bug if that makes sense I'm like thank you Ty I'm never gonna look at my nails the same again now it looks like two roaches but basically Vinny is uh pretty mad at me and uh yeah so were you guys able to find the landing zone by any chance the landing zone shirts because I don't know how to solve this I wanted to solve it today but by the time I finished all the feathered fun boxes like my back was out and it was like oh if I get done by this time like I could do all this other stuff and I just you know when you give up on the day you're like the day is over the day is over like it's over um somebody asked how to get your bird to eat veggies what I learned being a picky eater uh when we were doing our show parrot chop chef and I you know brought different veggies home that I never ever would have went in and like I don't know you know those weird ones like bok choy I don't know like weird things that like you know Megan you can't find the landing zone oh man um you know those you know those things like that well uh I would be surprised because like I also can't eat peppers and then we had peppers and then vinny ate them and I'm like what so it's really just about introducing and trying a variety another thing is that birds actually in the wild eat seasonally so that's why you could have like a whole bowl of like whatever let's just say seeds and um you you like uh you know there's a variety in there and then like three months they don't eat one seed and then suddenly the season changes and they do and I learned a lot about that by reading a book called the parrot who owns me she is I think she's an ornithologist and uh she she was used to working with wild birds and like observing them but she got an amazon parrot and so it's really interesting watching her apply the wild behavior to them um yeah I know okay hold on I gotta find this shirt this is like why I I try to explain to Ty sometimes I'm just not prepared for a live because I'm gonna say things that are not even ready yet because look my goal is never to come on and like you know try to sell something my goal is I'm excited about it I know you're gonna be excited about it and I want to tell you something about this shirt I called it the landing zone shirt Ty can you go to google like parrotstation.com and see if you can find this shirt because I don't want to go through your phone but um okay so the craziest thing one of you so I did a live yesterday and um actually it was very helpful because at the end I I was wearing the shirt so I had to tell you guys about it and um when I was wearing it uh I oh okay so this is why it's helpful so then I didn't realize that on the site we didn't have it that like you could order it internationally so of course and I get all these emails I think he wants that corn yeah he likes that I get all these emails uh saying hey I'm trying to order in Canada I'm trying to order it and one girl told me why she wanted to order it and it was because her bird passed and because the reserved sign is right on the shirt then she wanted it kind of like the memory of him sitting on her shoulder and I thought that was fascinating like the way energy moves because the way it transpires because that's why I created it my original concept was to have reserved for my bird George who had passed and um try the word hoodie and she and I wanted to have a rainbow at the bottom so you would know like this is reserved for the bird over the rainbow bridge because years after George passed I could still feel I'm sitting on my shoulder so um geez this is like a hidden damn link so I I um what I that's that was fascinating to me because that's why I had created it and I just thought like wow the feeling translated so wow this is really not coming up um I know where the link is it's can you go to YouTube and go to my um go to YouTube and go to my last live from yesterday that yeah now go to the pinned comments my pinned comment there's the link in there I made a short link click on it see if it works wow iPhone that's iPhone iPhone weird did that go to it what is it doing don't tell me yes that's it can you put that in the chat yeah for anyone that wants this by the way I think also I also didn't think this through but if you want the shirt and other toys or anything in the store just order them separately because it will hold up your order so what do you know what I'm saying like so just I just put it in the chat okay tie put it in the chat hopefully it doesn't do like you know blocked so that's my gym partner do you guys see the link or did it block him does it block you I don't know either maybe it'll let me see so orientation is locked okay let's see the messages it's a full link oh man and my head just had to text it to you yeah no it's not working because it's like okay if any of you I'll text it to you I'll text it to you and then you have to post it okay man well if any of you want to go into my live from yesterday it is pinned in the first comment but I don't know how to get it into this chat maybe it'll let me put it huh you're saying that you think it'll let me post it hold on right somebody might be right because in here in the thing is that the right link no that's not the right that's not the right link after all that time no no after everybody's looking up my nose can I sniff you can I sniff you she's looking not happy about that look watch always be prepped for them to move fast only I or wrenches can post it who's a wrench a moderator let me go back and go to your other tie you literally lost the link tie lost the link no I did it was right here it is in the pinned comment that's it but I did see it wrenches are my moderators wow I've been okay Wendy wants to give you a kiss okay this is a kiss from Wendy come here Wendy you literally just kissed Leo come here Leo okay now oh my god oh you sent it open oh I love this bird I'm obsessed with him so George told me hold on I'm gonna get you guys this link couldn't stop it you so cute and then it'll let me I have to do no I have to get I have to open it up is it gonna okay ready how do I write in there how do I write in there tie I don't even know how to write no that's my pen from my phone I can't even write in here literally I can't write in here you gotta be kidding me well I tried to write it and it didn't let me write it wow where is like the part where I can write how can I not be able to write to everybody share no wow that's crazy guys I don't even know pinned to live stream okay tag products but that's not it Leo's wants to I don't even know I'm literally gonna have to be sitting here going WW dot WWW dot okay no I can't even believe that I can't get you this information it is ridiculous I you know I'm gonna tell George we're gonna link it in a story and all that stuff oh my god first of all I don't know what I did it looks like it wants to put a filter and I don't know how to make it go away if you press that what if you end the live you're literally gonna end the live I'll sniff Leo for you I don't know what is that that's not the button that's literally the camera tie maybe we just choose one let's choose no filter and we'll be done I could do with a filter right now that's another you gotta be aware of where your face is in proximity to a bird that is in love with somebody else at the moment dude I ruined the this is I don't know what I did who else loves when their feathers are all messed up on top of their head like you look like so let's choose a filter maybe that'll help no you'd think I'd choose a filter and it would go away make this go away I can't I can't even give you guys a link I can't even see you anymore it's like it's like what is this I don't need like for real for real for real Ty what do we do here I think this is uh time to sign off oh my god I'm making people see sick I feel so bad I feel you I get like that I get like vertigo like that wow that's bad I'm sorry Peek-a-boo found the link Peek-a-boo's advanced Peek-a-boo's advanced Peek-a-boo's advanced well but I will tell you the link is listen we don't make things easy to purchase here okay Megan go to my old live the live I did yesterday and go to the first pin to comment you'll find it that you'll find it there there you go Peek-a-boo found that oh cockatiel so cockatiels are the only bird the main bird that emotional if I close my eyes you know how you think you can't imagine smell like you like I don't know you know how you can hear a song you can like have a memory but like you think you can't imagine smell like I can smell cockatiel sniff anytime I want in my head I don't know how but it's like what's wrong oh he's mad at Ty don't be mad when a bird is mad you probably should not put your face next to the bird no um but I take risks because I've been with them all my life you know so sometimes I do take risks I know he's mad he's jealous of this bird he doesn't care so much for Ty so much but the fact that there's another bird in the mix next to me is not making him happy he wants to get a close of it um but yeah then in that time see he's like he's gonna be grumpy and everything so you gotta be careful the way you interact with them in that moment you gotta make bird number one feel like bird number one my favorite bird of mine or just in general like picking one of your favorite kids I don't know what you do that well I love Leo the most um I like you could probably give me 10 moustache parakeets what oh he's got his eye on you by the way guys my doggy is like right down here and like just like he's super attached to me he follows me everywhere it's so adorable he's sleeping on your boot here I know he's so cute and um I don't know what I was gonna tell you but yeah moustache parakeets are I think like my favorite um but let's be more specific male moustache parakeets okay because Brando is the biggest brat you went down to the customer help watch me get customer service email tomorrow it goes to the trap master it goes to the trap master you need to fix your moustache and then it's all me I'm like okay I saw you on the live yesterday but yeah no um actually I did I got emails yesterday after my other live with like hello I can't how did they even find the link how did they find the link I don't know I have no idea oh my god a scarlet macaw how exciting you know I've never sat around and wanted a bird which is an interesting fact about me but I have friends that are like oh I need to do the bird I want this bird I want that bird I never want any bird can't get that bird I love them all but like I don't sit around saying I should get another one like that never happens they're not like Pokemon or anything then search for crop top or hoodie oh the rainbow bird crop tops are amazing hey good night love bird loft thank you so much for helping out with everything oh yeah the crop tops are kick so here's the thing they're not like up to here I don't walk around like that um if I could I probably would but I'm too cold anyway uh but I won't I mean I can't I don't let's just be real I'm inflammatory anyway so they're not like that they're really cool you could wear like some high-waisted jeans and everything really good like I was even what please answer my question I have to go to bed I don't know what your question is it went by so fast by the way on half of our screen is gone is there a parrot you have never seen how can I gain my bird's trust what parrot have I not seen in oh yeah some of the black cockatoos I've seen a black palm cockatoo I have not seen uh like uh yellow tails black cockatoo a red tails black cockatoo uh all the rest of the black cockatoo how to gain bird's trust is pretty much the the same way I've been talking about with like you know taking like go to a neutral territory that's not their cage something they're not territorial over and uh you know bond with them sit with them like you know if you're watching tv the bird's sitting with you watching tv even if they don't want to be on you let them be in the room like but away from their cage away from anything you know uh and preferably if you want to do this kind of work it's best alone without your partner unless your bird's not bonded to any of you then you want to socialize the bird with both of you so that the bird doesn't fall in love with just one of you because that happens all the time sometimes I'm like oh you want your daddy that's great because I already have 10 of you attached to me you know so um you know I'm a parent mom you know so you get exhausted and you you know it's real like these birds are with me for life so um right rocky rocky is a daddy lover I know but he's so cute and I think the main takeaway I would love for you guys to get from this video today is that they make sure that they don't have to bond with you make sure that even if you haven't come all the way or they haven't bonded with you yet or they're a little more wild that you don't say it's not working for me does that make sense like a lot of people are like oh they're not what I want them to be so you know don't come in like that with your your ideas and your shields of how they're supposed to be let them be who they are and work around it if if this bird never wants to step up to me I don't care like she can be free doing whatever and you know freedom is the most thing they need they need to be engaged and not caged that's really what they need to be you know as as much as possible honestly so okay Megan's kicking me off go rest I gotta go to the gym now not kidding we got like a gym partner pack my shirt Megan why don't you come visit me in my office okay come see what's going on Megan's the one that bluebird right she has yeah yeah she does she has she has uh also a green one now yeah yeah so yeah come by all right I love you guys so much Megan's kicking me off um she's paired station royalty so she knows what's what I want to thank you guys for the super chat thank you for joining in I really hope that you guys found the shirt clearly it's not promoted yet or we would have all that in order but those of you who are finding it you are in like you are like you're in the know like meaning like no one else knows and no one's else supposed to know yet I just feel like I'm talking with you guys like you know uh pack me a shirt with Megan well when I get off of this tomorrow I can't pack the shirts I don't have them yet remember it is uh it is pre-order so and not only is it pre-order but my printer I'm not letting him print anything when Mercury's in retrograde so even if I had all the orders like in the next five days which is not going to happen because I haven't even had time to like promote or whatever uh he he's not he's not he's not like I can't have any time he prints something when Mercury's in retrograde he's he's just he's frazzled so I want it perfect for you guys um name of the channel of the person next to me this is Ty Ty doesn't have a YouTube channel but he has a TikTok with his chameleon his chameleon's name is lifesaver and the channel is lifesaver the chameleon lifesaver the chameleon and you guys most gorgeous chameleons my nephew he's the most gorgeous chameleon you've ever seen in your life um ever ever ever Lori the crop tops you will love them they're so bright and they are so amazing and I noticed like there's not so much difference in length with the sizes more kind of like it makes it more flowy so even if you felt like you could wear a small you could also wear a large does that make sense but it it's not because it's not a tight crop top if that makes sense so however you feel comfortable um I don't know if that makes any sense yeah but they do they are bigger in size and smaller they are it's just I don't know anyway Megan said I better go yes go and click on that secret link I love you guys so much I will see you uh I'm gonna go live more and um uh thanks Ty for making me come out today and we will be back love you guys yeah you like that oh my god we're excited about now look at that he's excited bye Fatima you guys all the amazons wake up now they're like oh it's time to be alive well you're excited about you're so beautiful hi I'm obsessed with this bird hi bird all right I got I love you guys so much exactly I don't have to edit it but I do want to give you guys all our content that we do film she's got to get on it manifest me oh a production Josie was about to manifest that finger in her mouth Josie's like all right love you guys so much bye oh tell Annie tell your pug I loved him I assumed it's a he I don't know but I love you guys so much bye