Principles of neurobiology

Beique

Lecture 1

The origins of neuroscience

People found skulls that contained holes in them that had partially healed. The Egyptians thought that the brain wasn’t important; they would remove it from the nostrils. Hippocrates thought that brain was the seed of intelligence. Aristotle thought it was there just to cool the body. Galen was the doctor for the gladiators, made a bunch of analysis’ of gladiators who received blows to the head, studied sheep as well, came up with cerebrum, cerebellum and ventricles – he admitted the hypothesis that the cerebellum was involved with movement and that the cerebrum was involved with sensation and perception – he was right but his conclusions as to how he came to this were trivial. The Renaissance convinced that the ventricles and the nerves were little hydraulics. Descartes thought that everything that was human was human because of God. 17th and 18th century – found the white and grey matter, they thought that every brain structure had a specific function because they realized that the colours served different purposes. 19th century – could elicit muscle function with electrical currents, a biological phenomenon could be an electrical phenomenon – so they are not hydraulic systems. Bell and Magendie took the dorsal horn and thought that the roots leading out served different functions, they found that if you cut the ventral part it leads to muscle paralysis, if you cut the dorsal part, the animal is deprived of sensory information (muscle spasms). Flourens wanted to see what would happen if you removed certain parts of the brain. Gall idea that looking at the shape of a head you could determine personality traits – Flourens was totally against this, while he was right, he was also wrong by saying that “all regions participate equally”. Broca – brocas region related to learning. Gages story – involved in formation of explosives, it exploded in his face and a metal rod blew through his skull. Not only was he alive, he was also conscious, this was a perfect lesion in his prefrontal cortex which caused a personality switch, and he became aggressive and such. Natural selection is evident when looking at brains of our common ancestors, the brain has a historical remenance of evolution, it gives credence to the use of animal models. There is very little you can do with the human brain – ethical reasons – in modern science, there’s an inverse reason between what you want to study and what you can study. Thus we study animals. Golgi invented golgi staining in order to view neurons, Cajal wanted to disprove Golgi so copied his work but it worked so they both got the nobel prize. Kuffler and Katz believed that neurons were based on a chemical basis, however Eccles believed it was an electrical basis

Lecture 2

Nissl stain labels neurons and not glial cells, it labels DNA so the nucleus. Histology is the study of tissues. We take brain tissue, slice it up and insert the Nissl stain and it will stain the neurons. Where the area is darker, there is a higher density of neurons. When looking at the cortex with the Nissl stain, it is obvious that there are different layers (6). Cell bodies typically found in layer 5 and 6 and send dendrites all the way up to 1. Pyramidal cells are found throughout these

layers. Golgi stain will only stain certain cells, good to use when looking at the morphology. Apical dendrites are longer and go upwards. Basal dendrites go downwards. Axons have a constant diameter. Cajal hypothesized that neurons had an ability of plasticity. You can’t see synaptic clefts with light microscopy, but you can see is with electron microscopy because the resolution is better. You can use light microscopy to study the shape of neurons. Neurons are by far the most complicated cells in the body. The number and types of receptors that neurons have in their membrane determines what they do. Membrane proteins will be made in rough ER. Neurons have a huge energy requirement, the making of the gradient costs a lot of energy, high supply of mitochondria. Protein concentration differs depending where it is positioned. The cytoskeleton is not static, they are constantly moving, when you increase the activity, the neurons will increase in number. Axons are not linear; they can have axon collaterals which are protrusions of axons that are travelling to other regions of the body. One neuron can innervate many things. There is no ER in the axon. At a central synapse, you will have 0 to 1 vesicle release by action potential. Anterograde transport is from the nucleus to the terminal. Retrograde transport is from the terminal to the nucleus. Analogue signal is continuous in time. Digital signal is discontinuous in time. Neurons are converters, the dendrites receives all the information from other neurons and has to decide whether to shoot an action potential, this is an example of analogue signal. But the action potential itself is a digital signal, it is all or none, it carries information in time. Neurites are axons and dendrites. Spines are the site of excitatory transmission, but you can also have these transmissions on aspinous axons. Pyramidal cells in the somatosensory cortex and visual cortex are extremely similar. Interneurons are within the cortex but make contacts with their close neighbors, they almost always release GABA. Astrocytes are the main type of glial cells, there are pretty much found all over the brain. The glutamate synapse is the essential transformation of information in the brain. Glial cells participate in this; they activate receptors to clear the glutamate so that if another event enters, the body needs to differentiate between the 2. So glutamate is actively being cleared from synapses by glial cells. Myelin sheaths are made from oligodendrocytes and form concentric circles. Microglia act as phagocytic cells and function with the immune system.

Lecture 3 – the neuronal membrane at rest

**describe a membrane at a resting potential

One of the things that neurons do is that they transmit information via the neural code. This is action potential discharge. Neurons are resting at -60mV. The neural code is the amount of information that is being transferred and this will determine if it should fire an action potential or not. Water is a polar solvent; the ions are not evenly distributed. If we take NaCl and we let it dissolve in water, the affinity for the Cl- molecules has a higher affinity for positive ions (H) than the negative ions (O). In the membrane we have ion channels and they need to be able to differentiate between ions. They can only however recognize different ions when they are in contact with water. To make a transmembrane protein, 5 subunits come together (common) with a hole in the middle forming a pentamere. Protein channels contain distinct termini. Typically the N terminus will be found on the outside and the C terminus on the inside of the lipid bilayer. These channels open and close (gating). Channels are different than gates. Channels allow simple diffusion – passive. Whereas pumps need a concentration gradient – active, require

energy. Electricity – resistance is the same as conductance but they are the opposite. Batteries generate power across a resistance. The resistance through a bilayer is infinite. If we put holes in through the bilayer, the current will flow through these pumps. I=gV, if g and V increase then I increases as well, where g = resistance and V = voltage. If you stick an electrode in a neuron, the potential difference will typically give you a negative result, which means that one side of the membrane is more positive than the other side. We know that the inside is less positive (or negative) than the inside. The concentration on either side of the membrane is offset by the concentration of K. because the concentration of potassium is usually higher on the inside of the membrane, it will have the tendency to flow through its pump towards the outside (going down its concentration gradient) which will then make the inside of the membrane more negative. We always refer to an electrical chemical gradient because ions are charged. At this point, the membrane is at equilibrium and this is the resting membrane potential. But there is a constant competing force because the potassium will then want to flow into the cytosol. Equilibrium brings on the driving force which is determined by Vm-Eion. If your cell is at equilibrium potential for potassium (-80mV), and we open the potassium channels, the driving force will be 0 because there will be no flow. The equilibrium potential is determined by the charge and the concentration. If you have a combination of monovalent and divalent ions, this can cause an issue with the reversal potential. Know the terms of the Nernst equation. Have a rough idea of the reversal potential. So you have a high concentration of a positive charge (potassium) inside of the cell at a resting membrane potential. The resting membrane potential of the cell is going to go towards -80mV. For sodium, you have a higher concentration on the inside of the cell, if you have a pump that is more permeable towards sodium, then the resting membrane potential will be 62mV. Potassium channels are almost always open and sodium channels open for short periods of time. All of the behaviour of neurons is determined by the concentration of ions which can be determined with the Nernst equation. For calcium, there is less calcium than potassium outside the cell. Sodium and calcium have similar behaviours, when the channels open; the potential will want to go towards the reversal. For chloride, the concentration is higher on the inside than the outside (opposite of K, but similar to Ca and Na) but the reversal potential is similar to K – this is because Cl is an anion. The sodium-potassium pumps are slower than the ion channels and they require energy. 70% of the brains consumption of ATP is due to these pumps. They pump 3 Na ions out and 2 K ions in at the same time. So when that sodium pump is open for that split second, pushing sodium out, this also cause potassium to enter. When potassium channels are open, the conductance is high. Sodium at rest is 0. A depolarization occurs when there is added potassium on the outside of the cell – it gets less negative. If you give someone an excessive amount of potassium, this will cause a depolarization and this will kill someone. Glial cells (astrocytes) buffer changes in potassium.

Lecture 4 – action potential

Hodgkin and Huxley had access to squids in which they could use to study electrophysiology. The giant squid has a giant reflex axon, and bc they were so big, they were experimentally favorable. The studies they performed on the squid ended up being very similar to the human. These guys built oscilloscopes to view action potentials. An action potential starts off when the neuron is depolarized, but the neuron starts off in its resting potential. The depolarization is caused by a current, once, and only when it meets its threshold, will an action potential occur.

Action potentials are non-linear. Once the depolarization reaches its threshold, an overshoot will occur, which is an all or none result. Following this, there will be undershoot which will fall underneath the resting membrane potential. This process takes about 3ms to 1ms. To view action potentials artificially, we can insert electrodes which inject current. Direct current injection will cause several action potentials. If we were to block sodium channels, the neurons will act more like a passive structure (similar to injecting current in a blob of lipid), therefore there won’t be any depolarization beyond threshold, and therefore there will be no action potentials. Voltage dependant channels are active conductors. Firing frequency reflects the magnitude of the depolarizing agent. So if we add a little bit of current, threshold won’t be reached so it will plateau at around -70mV. The more current you inject, the cell will give rise to a linear firing action potential. Membrane and conductance can be used interchangeably. Using the Nernst equation, g determines the conductance. If the conductance for potassium is 0, there will be no action potential. If we open up gk, we want to know how much potassium will flow in; this is determined by the conductance and the driving force. At rest, your conductance for potassium is much higher than sodium. Once there is depolarization, the conductance for sodium surpasses the conductance for potassium. Once it reaches the peak, we have mechanisms that will bring it down, this is necessary because the information needs to be transferred very quickly. The mechanism used is the opening of potassium channels (differ from resting potential), the membrane potential will want to go to equilibrium of potassium which is -80mV therefore it will drop. Eventually the sodium channels will close which will cause the hyperpolarization or undershoot. Some of the potassium channels will close and we will go back to resting membrane potential. When putting an electrode in a neuron, you have 2 options, voltage clamp or current clamp. For voltage clamp, the amp clamps the cell at a certain voltage and keeps it there. So if you clamp it a -40mV, then all the sodium channels will open, but the clamp needs to do something in order to keep it at -40mV. Looking at the structure of the ion channels, they contain pore loops which confer specificity. Somehow, in their structure, there is a voltage sensor that is able to transduce potential difference in some sort of physical movement in order for the charges to go through. The voltage sensor is embedded in the membrane so that when there is charge accumulation around the membrane, there is a tiny movement by S4 which allows the ions to pass through. The membrane of the channels are sensitive to the size of the hydrated ions (K is bigger than Na). When studying these channels, you can insert electrodes and pick the conductance and observe the behaviour of these channels. When you put the conductance to -40mV, the sodium channels open for a very brief period of time, but do not open again because they are gated and will thus be blocked. This is due to the fact that there is a refractory period, so during depolarization, they cannot be re depolarized. There are a variety of toxins implicated in voltage-gated sodium channels. For example, a toxin coming from a buttercup, aconitine, lowers the activation threshold. Because this is more easily obtained, more action potentials will fire. The pump is responsible for making the Ek at -80mV at resting potential. Once the action potential has reached its peak, the potassium channels open in response to the depolarization, as do sodium, however they are delayed. The use of this is to rectify the membrane – it wants to go back to equilibrium. The absolute refractory period occurs when sodium channels are inactivated, they are gated. The relative refractory period occurs during undershoot phase. During undershoot, it is more difficult to reach the firing potential threshold which is the relative refractory period. When sodium channels open and sodium

rushes in, this is current. Potassium channels open longer and slower than sodium. So K is responsible for rectifying the action potential.

Lecture 5

L5 – Synaptic Transmission

Synaptic transmission• Information transfer at a synapse• 1897 – Charles Sherrington – coined term synapse• Chemical and electrical synapse

o 1921 – Otto Loewi – Vagusstoff – acetylcholine; saline from one frog heart preparation slowed another heart in another batho 1959 – Furshpan and Potter – first example of a clear electrical synapse in a crayfish

Types of Synapses• Direction of information flow

o One direction – neuron to target cello First neuron – presynaptic neurono Target cell – postsynaptic neuron

• Electrical synapses – very fast transmission• Chemical synapses – majority of synapses in brain

o Somewhat slower, generates postsynaptic potentials (PSPs)o Synaptic integration – several PSPs occurring simultaneously to excite a neuron (causes action potential)

Electrical synapses• Gap junction

o Channel – 1 connexon from both cells form the channel for ions to pass through

Connexon – formed by 6 connexins• Cells are said to be electrically couples

o Flow of ions from cytoplasm to cytoplasm – no delay in time when measuring electrical synapses between cells

Chemical synapses

The neuromuscular junction (NMJ)• Studies of NMJ established principles of synaptic transmission – multiple synapses off of axon attaching to muscle

Soup vs. spark – Katz – basic principles of synaptic transmission

CNS synapses (examples)• Axodendritic – axon to dendrite – primary synapses • Axosomatic – axon to cell body – mainly inhibitory synapses • Axoaxonic – axon to axon – can modulate excitability of axons • Dendrodendritic – dendrite to dendrite – rare and specialized • Grays type I – asymmetrical, excitatory – postsynaptic terminal is thicker then presynaptic • Grays type II – symmetrical, inhibitory

Chemical synapse structure:• Post synaptic density – receptors filled with scaffolding proteins between pre and post synaptic terminalso Contains the neurotransmitter receptors which convert the intercellular

chemical signal (neurotransmitter) into an intracellular signal (action potential) in the postsynaptic cello On glutamate excitatory synapses – grays type I

Principles of chemical synaptic transmission• Basic steps

o Neurotransmitter synthesiso Load neurotransmitter into synaptic vesicle – takes energy o Vesicle fuse to presynaptic terminalo Neurotransmitters spills into synaptic cleft – following a signal o Binds to postsynaptic receptorso Biochemical/ electrical response elicited in postsynaptic cello Removal of neurotransmitter from synaptic cleft

• Neurotransmitterso Amino acids – small organic molecules – glutamate (excitatory), glycine (mainly inhibitory in spinal cord), GABA (inhibitory) – not used to make proteins so evolution used them to make neurotransmitters o Amine – small organic molecules – dopamine, acetylcholine, histamineo Peptides – short amino acid chains (proteins) stored in and released from secretory granules – dynorphin, enkephalins

• Neurotransmitter synthesis and storageo Amines, amino acids, peptides – need to be synthesized and stored – happens in vesicle

Precursor molecule with synthetic enzymes make neurotransmitter and package them in vesicles which are waiting and ready fro release

• Neurotransmitter releaseo Exocytosis – process by which vesicles release their contents

o Mechanism Process of exocytosis stimulated by intracellular calcium – calcium rushing in Proteins alter conformation with influx of calcium– activates lipid vesicles to attach to pre synaptic terminal membrane Vesicle membrane incorporates into presynaptic membrane Neurotransmitter released Vesicle membrane recovered by endocytosis -

Calcium and presynaptic release – Katz and Miledi• Nerve terminal and axon from motor neuron (NMJ in frog) – obtain recording from muscle cell, stimulate axon with electrode and put in certain concentrations of ions into muscle• Bath – solution with no calcium • Stimulate axon nothing happen• Just before pulse insert solution containing calcium – then stimulate and get depolarization• Calcium after – nothing happens• Conclude that calcium is required for synaptic transmission

Calcium and presynaptic release – Llinas and Heuser 1977 • Obtain recording from presynaptic terminals and show presence when depolarize could elicit currents that were mediated by voltage dependent calcium channels• Without electrical stimulate calcium alone could cause depolarization

Stochastic nature of neurotransmitter release • Release probability – when have action potential that reaches terminal more often then not nothing happens • Has to do with how many vesicle are actually ready to be released • High computational requirement – release probability of 1 – release all the time • Fatt and Kats – stimulate axon electrically there is a delay while vesicles are joining and releasing, then see responses

Principle of synaptic transmission • Quantal analysis of EPSPs – excitatory postsynaptic potential

o Synaptic vesicles – elementary units of synaptic transmissiono Quantum – an indivisible unito Miniature postsynaptic potential – “mini” – one vesicle being released o Quantal analysis – used to determine number of vesicles that release during neurotransmission

Spontaneous – Count blips and calculate amplitude to see normal distribution

Electrical stimulation – Plot amplitude and found both had exactly same amplitude and spontaneous ones – all peaks are position at a multiple of 2 of the previous peak This lead then to hypothesize that we have synaptic release but its all a multiple of 1 – a quantum – something akin to a vesicle being released, the more vesicles the higher the amplitude

o Neurotransmitter junction – about 200 synaptic vesicles, EPSP of 40mV or moreo CNS synapse – single vesicle, EPSE of few tenths of mV o Different release probabilities for different synapses – changes fundamental computational release of vesicles o Release probability changes by:

Change ability of vesicle to bind to membrane Or reduce conductance of calcium channels

Principles of chemical synaptic transmission• Neurotransmitter receptors

o Ionotropic – transmitter-gates ion channels • Excitatory and inhibitory postsynaptic potentials• Excitatory PSP – transient postsynaptic membrane depolarization by presynaptic release of neurotransmitter• Inhibitory PSP – transient hyperpolarization of postsynaptic membrane potential caused by presynaptic release of neurotransmitter• Neurotransmitter recovery and degradation – important for point transmission, targeting specific axons

o Diffusion – away from the synapse – lower concentration that farther away, so synapses react where there is the highest concentration of neurotransmitters o Reuptake – neurotransmitter re-enter presynaptic axon terminalo Enzymatic destruction inside terminal cytosol or synaptic clefto Desensitization – ex. AChE cleaves Ach to inactive state

Reversal potential of EPSPs – same principles that drive ion flow through voltage-gated channels

• Ligand gated channels – gated by ligand • Nonselective – permeable to both sodium and potassium – reverse at 0mV

Neuropharmacology• Effect of drugs on nervous system tissue• Receptor antagonists – inhibitors of neurotransmitter receptors – Curare• Receptor agonists – mimic actions of natural occurring neurotransmitters – Nicotine

• Defective neurotransmission – root cause of neurological and psychiatric disorders

Principles of synaptic integration• Process by which multiple synaptic potentials combine within one postsynaptic neuron – neurons are a-d converters – need multiple signals to build up to an action potential – summation of multiple signals is an action potential • Determining length of EPSP – opening of receptors , length of time in open conformation, passive electrical properties of cell• EPSP Summation

o Allows for neuron to perform sophisticated computation – contingency rules o Integration – EPSPs added together to produce significant postsynaptic depolarization – integration over time and space o Spatial – EPSP generated simultaneously in different spaces – separate action potentialso Temporal – EPSP generated at same synapse in rapid succession - same axon eliciting multiple action potentials right after the other

• The contribution of dendritic properties to synaptic integrationo Dendrite as a straight cableo Membrane depolarization falls off exponentially with increasing distance – input at distal dendrite input needs to make it to cell body – if only follow cable properties chances are the input wont make it there

Vx = Vo / e^x/ o Dendritic length constant (λ)o In reality, dendrites are very elaborate structures that contribute to more complex integrative properties 

• Excitable dendrites o Dendrites of neurons have voltage­gated sodium, calcium, and potassium channels (active properties)

Can act as amplifiers (vs. passive) Na channels – Different in type and density (low) – boost signals 

o Dendritic sodium channels – may carry electrical signals in opposite direction, from soma outward along dendrites 

• Inhibitiono Action of synapses to take membrane potential away from action potential thresholdo Exerts powerful control over neuron output 

• IPSPs and shunting inhibitiono Excitatory vs. inhibitory synapses – bind different neurotransmitter, allow different ions to pass through channelso Membrane potential less negative than ­65mV = hyperpolarizing IPSP

• Shunting inhibition – inhibition current flow from soma to axon hillock – poking hole in hose

Concluding remarks• Chemical synaptic transmission

o Rich diversity allows for complex behavioro Provides explanations for drug effectso Defective transmission is the basis for many neurological and psychiatric disorderso Key to understanding the neural basis of learning and memory

Spatial summation is when you have inputs that come into the cell simultaneously. So all the action potentials are added up and the body doesn’t know exactly how many action potentials there were. With temporal summation, there are many synaptic inputs that are one after the other and will not be added up but will continue to increase the peak of the action potential. If you inject a current in the cell, the electronic properties will decrease over time and fairly quickly. So we need to find a way so that the inputs can be transmitted to the cell body. Dendrites also have active properties such as voltage gated channels whereas before we thought that these were passive. When we talk about density of the sodium channels in axons, it’s very high. However in dendrites, it isn’t as high. Think of the principle of shunting inhibition as a soccer ball with holes in it. So if you conduct an EPSP, and you measure at that point, you will get a reading with high amplitude, and then if you measure it at the body, this amplitude will have decreased and the peak will have also expand. If you conduct the same experiment, but near the cell body you conduct a IPSP, once you measure the conductance of the cell body, you will get nothing. So basically, this shunting is like poking holes where you can keep putting air in your soccer ball, but it will still not get bigger.

Lecture 6

The peptide neurotransmitters aren’t as active in synapse as the amines. Fast transmission is mediated by ligand gated channels (ions channels) and slow transmission is mediated by g protein coupled receptors. Some NT will trigger activation of ion channels. To study nt, we have to find where that specific nt is synthesized and stored: to do this, we can do immunocytochemistry or in situ hybridization which localizes synthesis of protein or peptide to a cell – for example, we know that acetylcholine is made from tryptophan, so we can track the enzyme that makes tryptophan and we can detect the mRNA to map proteins. We also need to study neurotransmitter systems, we take brain tissue from an animal, and we can stimulate synapses on this live tissue. And finally we need to study synaptic mimicry, this is when there is a molecule that evokes the same response as the nt (agonist). So we can take glutamate and locally stimulate the neuron and observe the response and then take another nt to see if it has the same effect. Krnjevic is one of the first ppl to find that some amino acids can be nt, like glutamate. Glutamate will activate many receptors that are not necessarily of the same origin. You can have a cell with a serotonin receptor that inhibits cyclicAMP, but you have other serotonin receptors that will activate cAMP, so some drugs will actually activate both of these at the same time. ACh receptors can be different such as nicotinic and muscarinic receptors. Glutamate activate AMPA, NMDA, and kainite receptors. Looking at receptor protein classes, we have direct gating – which is ligand gated which is a channel. We also have indirect gating –

which is also ligand gated, but g-protein coupled and is not a channel, it is a metabotropic glutamate receptor, glutamate will bind to this protein which activates downstream protein signaling and will go on to do many other things. Dale’s principal depicts that with one neuron, we have one nt, this is only partly true. For co-transmitters, two or more transmitters can be released from one neuron. Neurons need to make nt, and sometimes this involved complicated machineries. We can use in situ hybridization to view the cholinergernic neurons. Other nt candidates and intercellular messengers such as endocannabinoids release glutamate and GABA when stimulated and this is directly related to hunger, hence the munchies when you’re high. Ligand-gated channels are usually formed from 5 subunits, this dictates key biochemical properties. If you take 2 different receptors that you think they would show a very tiny difference and you put them in an elaborate system or realistic models, then these small differences can have a huge effect on how the system functions. Nt have very similar properties – we won’t reinvent the wheel twice. They all have the same function – they bind to receptors. But it’s which receptors they bind to which make the difference. On the outside of the cell, a complex protein structure is present in which the binding site for the nt is present. Amino acid gated channels include glutamate gated channels which are permeable to both sodium and calcium. Calcium is super important because it is well compartmentalized. So when it is present in one synapse, the other synapses won’t see it. Turns out that the selectivity for calcium is … it acts as a buffer for the brain. Is calcium is a messenger, then we know it’s going to activate “stuff”, so every protein that are sensitive to calcium, you would think that calcium sensors would have high affinity for calcium, but they don’t. so the receptors are only activated when calcium is incredibly close – this is where the selectivity comes from. The most fundamental aspect of synaptic plasticity is the ability of the synapse to detect coincidences – molecule responsible for this is MNDA receptors. These are gated by glutamate, so when they open, they are permeable to K, Na, and Ca. when glutamate binds to these receptors, the gate will not open. But when your cell becomes depolarized (-30mV), then the Mg gets relieved and then there is outflow and inflow of ions. We also have GABA a receptors, which are permeable to Cl, they participate in inhibition and they reduce the excitability of networks that have gone haywire. They also have binding sites for a number of molecules such as barbiturates, ethanol and benzodiazepine. GABA is an antagonist; there are 2 different kinds of antagonist – competitive and non-competitive. Competitive antagonists bind to the receptors and block the response and will compete with agonists for other receptors to thus block more receptors. G-protein coupled receptors are single polypeptides with seven membrane-spanning alpha-helices. When the nt binds to these, there is a slight movement in the configuration which activates the G proteins in which they are bound to. Monoamines, such as ACh, preferentially bind to g-proteins which will either have a stimulatory or inhibitory effect on cAMP. The mechanism for this goes as follows: when GDP is bound to the alpha-beta-gamma subunit, it is inactive. When GDP gets phosphorylated, this causes the separation of the alpha subunit from the beta-gamma subunit. The alpha s-u will activate effector protein 1 and the beta-gamma s-u will activate effector protein 2. **Know the 5 steps on slide 34**. The alpha GTP will typically react on enzymes. The beta-gamma will dissociate and acts on ion channels such as a special type of potassium channel called GIRK channel. If you activate the 5HGI receptor, the b-y s-u will activate GIRK which will cause the channel to open and thus will cause a hyperpolarization of the cell. Now the alpha is going to activate or inhibit other things, such as adenylyl cyclase. Alpha 2 will activate Gi(inhibitory), beta

are stimulatory (Gs) – sometimes these are both active so we’ll have a push pull effect. GQ are active when hallucinogens are taken. PK puts phosphates on a whole bunch of proteins

Lecture 7

What’s the basic behaviour that you can monitor that will allow you to understand the basis of learning and memory? Habituation, the animal has learned to ignore a stimulus that has no meaning. Associative learning includes classical conditioning as well as instrumental conditioning. The instrumental approach was an experiment done with Edward Thorndike. This is more complicated than classical conditioning because it requires higher level processing. You learn to associate a response with a meaningful stimulus (reward lever). There are several advantages in using invertebrate nervous systems such as that they have small nervous systems which don’t contain very many neurons, the neurons are larger in size, and they have small genetic codes. These organisms are very capable of showing habituation. Repeated electrical stimulation of a sensory neuron leads to a progressively smaller EPSP in the postsynaptic neuron. Every step in the process could be affected during habituation. This means that the synapses may have become weaker, the ability of the stretch over habituation may have diminished, and the postsynaptic neuron on the muscle cell may also not function as well. The cerebellum contains Purkenje cells which are activated by a number of axons. The climbing fibers send the axons on the dendrites of these purkenje cells. Also, we have granule cells which send axons on the purkenje cells. We also have parallel fibers on the surface of the cerebellum (know this image – slide 13). Purkenje cells have huge dendrites and they receive dendrites from parallel and climbing fibers. We stimulate the parallel fibers and take note of them. The conditioning (learning) is given by stimulating the climbing fibers with the parallel fibers for a short period of time. And then we stimulate the parallel fibers alone for a second time and we will see small values for the cells response – weakening synapses. LTD only occurs in parallel fiber synapses that are active at the same time as the climbing fibers. The synapses have stored something – coding of information. The memory is the change in synaptic strength. Induction (learning) vs. expression (memory). To make neurons weaker or stronger, we can increase release probability, we can also manipulate their sensitivity. So we stimulate parallel fibers, which activate the AMPA receptors which is our initial strength response. When the climbing fiber is activated, it allows the increase on calcium flow towards the inside the axon which increased the activity of PKC which will phosphorylate a protein which will in turn reduce the affinity of the AMPA receptor. So once you stimulate the parallel fiber a second time, the AMPA receptor will have a lower affinity for the nt which will slow down the reaction. This whole mechanism is called long-term depression. In the hippocampus, looking at the image on slide 18, we see an increase in strength of the neurons through long term potentiation. If we take the recording of the CA1 synapse, it releases glutamate in the cells and we can monitor their response. So if you stimulate inputs (blue) with an EPSP, over time these will become very strong and will stay like this for several months. If you take a second input and stimulate it (red)...? if you have 100 axons and we stimulate all of them and we increase release probability to 0.5 then we will induce release from 50% of the axons. The expression mechanism for LTP is… ? The glutamate that activates AMPA receptors will activate NMDA receptors once the summation of the depolarization increased from a high frequency stimulus – your post synaptic spine starts to get loaded with calcium which is not normal. So glutamate release and the

postsynaptic depolarization (from AMPA receptors) together work as the coincidence signal. The increase in calcium allows more AMPA receptors to be loaded into the spine which will in turn make the synapse stronger. This is bidirectional.