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Lecture Notes for Chapters 1- 6

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Lecture Notes for Chapters 1- 6 Bil 268

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These are the lecture notes for all the chapters from 1 - 6, excluding 7, as he hasn't gone over it yet! I will be including that and everything that he goes over in the review class :)
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Date Created: 02/03/15
BIL 268 01132015 Lecture 1 Introduction I What is Neurobiology It is the study of molecular organization of neurons and the ways that neurons are organized through synapses into functional circuits that process information and mediate behavior ll Why Do We Study Neurobiology To understand ourselves and to cure neural and mental diseases Alzheimer39s Disease dementia affects over 3 million Americans 90 billionyr Schizophrenia affects several million Americans 130 billionyr Stroke 3rol leading cause of death in US 25 billionyr Ill Analysis and Method There are different levels of analysis 0 Molecular Neurobiology Cellular Neurobiology Systems Neurobiology 0 Can be auditory systems receptor cells in the ear that are connected by the auditory neurons that send information to the brain This is the whole system from the receptor cells all the way to the brain 0 Behavioral Neurobiology Neuroethology o Neurocircuitary in the CNS to mediate certain behaviors 0 An example is the barn owl which can catch the rodent in total darkness due to a super sound location capability which means they can hear the sound from the rodents Cognitive Neurobiology o Belongs to the third part of the course 0 What is motion attention brainwaves etc General Scientific Method Observation 0 Made during experiments designed to test a hypothesis 0 Replication 0 Seeing if you can get the same results after conducting the experiment a number of times 0 Repeated on different subjects as many times as required to eliminate the possibility of errorresults occurring by chance 0 Interpretation 0 Verification o Other researchers will repeat your experiment to see if they are able to derive similar results 0 Used as a way to eliminate frauds IV Animal Care and Use Animal Welfare The Law The AWA in 1966 to protect certain animals from inhumane treatment and neglect Who Monitors the Use of Animals in Research The USDA A pus monitors this Animals are all warm blooded species except for birds rats and mice and farm animals used for production Majority of research animals gt90 are rates and mice USDA monitors the use of less than 10 of animals primarily pigs rabbits guinea pigs hamsters goats sheep cattle horses dogs and cats Who Oversees the Use of All Animals Especially Rodents Public Health Service does Requires institutions receiving federal funding to submit an assurance statement to the OLAW Office of Laboratory Animal Welfare States that the institution is committed to following the Guide for the Care and Use of Laboratory Animals The Guide 8th Edition Provides details as to the type of housing veterinary medical care for animals Also defines how a designated committee must review animal care and use protocols Animal is defined as quotany vertebrate animal used in research testing or teaching IACUC Institution ofAnimaI Care and Use Committee Procedures to Monitor Animal Care and Use IACUC must inspect and evaluate its facilities and programs at least twice a year The USDA inspects covered species annually and more frequently if there is an issue or a complaint AAALAC Association for Assessment and Accreditation of Laboratory Animal Care International Representatives inspect accredited facilities every 3 years IACUC Committee that oversees and evaluates the institutions animal program procedures and facilities It reviews all protocols describing animal use procedures may not begin until protocols are approved Derives their authority from the law USDACLAW It must include a minimum of 3 people Refer to ppt The Three R39s Reduction Use only as many animals as you need for an experiment and do what you can to reduce that number Don t use more than what you need Reduce the pain that you are causing the animals via anesthesia or drugs Refinement Improve your experiment using new techniques Replacement See if you can use models or techniques in place of live animals Animal Models Nematodes Insects squid zebra fish rodents monkeys Do not confused animal rights with animal welfare BIL 268 01152015 Lecture 2 Neurons and Glia Neurons are excitable cells There are almost a trillion cells in the brain There are more neuron cells than glial cells There are ten times more neuron cells than glial cells If you re looking at a typical cell what is the size of it Around 001 005mm in diameter I The Neuron Doctrine Histology the microscopic study of the structure of tissue There are five steps Fixation fixative such as formaldehyde paraformaldehyde or glutaraldehyde Section microtome Stain Nissl stain to stain cell bodies Mounting 0 Examination using microscope Confocal uses a laser and depending on the wavelength it can go really deep Using cutting edge of microscope you can see the structure of cells but you cannot distinguish between the neurons and glial cells Nissl stain is used for this 0 Chemicals are used to distinguish the Nissl bodies Rough ER 0 Neurons have a Nissl body glial cells do not Neurites term for dendrites and axons Tubes that radiate away from the cell body Camillo Golgi came up with the Golgi stain Cutting the brain into small pieces and soaking them in a solution of potassium dichromate and after a couple of days you put it in silver nitrate cover and protect from light for a couple of days The first solution impregnates the tissue and so does the second solution and a chemical reaction is generated which creates a precipitate 0 Color range from brown to red Brown labels are for neurons The neurons are stained entirely and you can see the cell body and the neurites In a neuron you can clearly find the axons and the dendrites The dendrites are branched and are shorter than the axons Dendrites have a larger diameter than the axons Dendrites function like an antennae and they receive input to the neuron and the cell bodies There are tiny bumps on dendrites which have the function of receiving signals and are also where you can find polyribosomes Multiple dendrites but only one axon o Axon can be divided into many different branches Golgi proposed a reticular theory a neural net He also believed that the neurites were fused together for different cells ultimately proven to be untrue Golgi type I vs Golgi type II neurons Golgi type I are neurons with long axons that extend from one part of the brain to the other Golgi type II are neurons with short axons that do not extend beyond the vicinity of the cell body Cajal used the Golgi stain method to conclude that the neurons were very close and were not fused with one another He also proposed that neurons are polarized in terms of morphology and function The Neuron Doctrine neurons are the elementary functional unit of the nervous system and they communicate with each other via contact and not continuity Brainbow is when you39re looking at the neurons inside the brain which have different colors and are based on fluorescent proteins Expresses the foreign genes not originally in the host gene Confocal microscopy differs from the traditional microscope in the sense that it uses lasen Connectomics A new field in neuroscience to study the map of neural connections Exploring the connections down to the smallest level and they also want to look at the whole brain II The Prototypical Neuron Know what the cytosol is and what the cytoplasm is The DNA is in the nucleus Whenever you have different functions you have certain types of proteins The Nucleus Includes the double stranded DNA which is first separated using one strand as a template Breakthrough of 2007 Human Genetic Variation The human genome project took a decade Everybody s genomes are different which is what makes them different from each other Changes in the number and order of genes add variety to the human genome The Soma It is the place for protein synthesis Protein synthesis at the rough ER inserts into the membrane and becomes membrane proteins Know the DNA where you can find it Transcription and translation know the process The Neuronal Membrane It is a phospholipid bilayer with a hydrophilic head and a hydrophobic tail Membrane proteins can be found via ion channels and pumps Pumps require ATP in order to function and it pumps ions against the concentration gradient from a low concentration gradient to a high concentration gradient 0 The function of the pump is to maintain the concentration gradient of different kinds of ions across the cell membrane on channels open and close allowing ions to move 0 The ionic movement is always from high concentration site to low concentration site via diffusion The Cytoskeleton Made up of microtubules neurofilaments and microfilaments They give the neuron its shape Tubulin is a small structure made of smaller strands Polymerization is the process of joining small proteins to form a long strand and it results in a polymer MAP s are a class of proteins that participate in the regulation of microtubule assembly and function Axon The axon begins at the cell body in a region known as the axon hillock The spike initiation zone is depolarization of the cell membrane Action potential travels to the terminal of the axon Moving away from the cell body is the efferent fiber Other neurons that are coming towards the cell body to communicate are afferent fibers There is no protein synthesis in the axon Branches of the axon are known as axon collaterals Information is sent down the terminal of the axon and is called the transport axoplasmic transport Direction can be transport down the axon to the cell body or the opposite Kinesin is related to the anterograde transport Dynein is the protein that allows for retrograde transport Synapse Communication between two neurons is done via the synapse Mechanical synapse is the communication that uses the chemicals of the neurotransmitter Know the basic structures There is a gap between the presynaptic and postsynaptic membrane known as the synaptic cleft Ill Neuron Classification Based on the number of neurons Can be unipolar bipolar or multipolar Most neurons in the brain are multipolar They can also be based on the shape of the dendrite Pyramidal Stellate Spiny and Smooth You can also identify them based on the connections the length or even the neurotransmitters IV Glia There are different types of glial cells Astrocytes they remove neurotransmitters from the synaptic cleft they control the extracellular ion concentration and help determine the growth pattern for neurites Myelinating Glia There are two types Oligodendroglial and Schwann cells Schwann cells are only found in the PNS while oligodendroglial cells are found in the CNS The node of Ranvier is the region where the axonal membrane is exposed BIL 268 01202015 IV Glia Myelinating Glia There are two types Oligodendroglial and Schwann cells Schwann cells are only found in the PNS while oligodendroglial cells are found in the CNS The node of Ranvier is the region where the axonal membrane is exposed The action potential needs to jump from one node to the other Other Glia Ependymal cells neurons that start in one area and have to migrate to the target structure with the guide of ependymal cells Microglia when cells die microglia remove the debris Lecture 3 Resting Potential There will be questions from the first exam that will pay attention to this section The major chemical players are Potassium Sodium Chloride Calcium For the RMP Potassium and Sodium are were important I Chemicals Cytosol and Extracellular Fluid Made up of water polar molecules ons cations and anions that are monovalent and divalent Cell Membrane Made up of a phospholipid bilayer The head is polar and hydrophilic and is made up of phosphates The tails are non polar and are hydrophobic and are made up of hydrocarbons Protein Protein is composed of amino acids and each amino acid is made up of an amino group a carboxyl group carbon hydrogen and a residue group If the residue group is hydrophilic it can be inside or outside in the water solution or it can be hydrophobic it can enter the internal layer of the cellular membrane The primary structure of an amino acid is called a polypeptide The alpha helixes are put together to form a sort of a 3 d structure known as a subunit tertiary Several subunits put together form ion channels The ion channels allow ions that are moving in and out of the cell Some ion channels are open all the time Other type of ion channels are gated and have some conditions that is required to open them 0 The membrane has to be depolarized to reach a certain voltage and then open the channel I Voltage gated ion channel The ion channel can also be opened by mechanical stimulation 0 An example of is a cut on skin receptors on the skin got activated by the stimulation and caused the ion channel to open up Channel Proteins You need to know that there are three important properties about ion channels The first is called ion selectivity ie Na ion channel or K ion channel 0 The ion channels primarily only allow certain types of ion passing through the channel The second is gated ion channel The third is when ion channels are open ionic movement always goes down along the concentration gradient 0 That means always moving by diffusion from high concentration side to low concentration side 0 Also the membrane protein or the enzyme The major difference between the pump and the channel is looking at the ionic movement ll Movement of Ions Diffusion Diffusion refers to the movement of ions from regions of higher concentration to regions of low concentrations Net movement is always from left to right till you reach equilibrium Selectively Permeable Membrane means that the membrane is open only to a speci c kind of ion and the other ions are shut out This membrane potential difference is generated by ion movement When you have a membrane potential you have two different forces together Electricity Ohms law states that the current is equal to the voltage divides by the resistance If your breaker contains a 1M NaCl solution and you have a battery with a negative and a positive pole The negative charge Cl is attracted to the anode and the positive charge Na is attracted to the cathode Ill The Ionic Basis of RP Membrane Potential VM The recording electrode and the reference electrode ground electrode are used to measure the resting membrane potential The inside of a neuron is more negatively charged than the outside 65 mV What is the process of amplifying You tried to record RP because they are usually pretty big 80 mV The tip of the electrode is very small and resistance is very high which means that when current is passed through the electrode a huge voltage drop is observed When you finally measure on the screen the measurement is one that has already suffered a big voltage drop You want to have the amplifiers resistance to be higher than the resistance of the electrode so that what is shown on the monitor will be very close to what you tried to measure in the RP Equilibrium Potential The equilibrium potential refers to the given ion You only consider one Or you can see the EP for Sodium or Potassium or Chlorine Right side is the extracellular side and the left side is the intracellular side and the ionic concentration is different In the membrane only selectively membrane to potassium and not the other ions Potassium moves from the IC side to the EC side which is why the positive charge moves outside and causes the membrane to become more positive Starting with diffusional force potassium moves from left to right until the diffusion force drops and the two sides have reached equilibrium What s the difference between equilibrium potential and resting membrane potential EP is for potassium ions and RMP takes into account all the other ions K Na Cl and Ca The EP for K 80 mV and the RMP is 65 mV Resting ion channels are open all the time and allow K to move from the inside to the outside Na can leak into the cell via the resting ion channel A positive charge going in increases the membrane potential till it finally reaches equilibrium Trying to measure EP for Na is 62 mV Once equilibrium is reached There is no net movement There are small changes in ionic concentration that cause large change in membrane potential The membrane stores electrical charge IC is negatively charged and EC is positively charged The ionic driving force is the membrane potential minus the equilibrium potential The Nernst equation can be used to calculate the equilibrium potential The Nernst Equation Calculate Eion Known the equation not the one from the book but the one from the ppt Refer to the ppt and read the slide Make sure you known the signs For Na it is 1 and for CI it is 1 You need to know the charge for a given type of ion You need to know the intracellular concentration and the extracellular concentration The unit is in volts for the charge of the ion Eion If the extracellular concentration if 10 mM and the intracellular concentration is 100 mM with a z of 1 and a T of 37 C what is the Eion Ion distribution across the membrane The ion pump is an enzyme You basically try to move the three sodium out while trying to bring the two potassium in which is why there is a drop noted in the graph Relative Ion Permeability The Nernst equation calculates EP The Goldman equation know in which situation to use which equation Know how to use the calculation for the Nernst equation Regulation of extra K concentration How does extracellular potassium concentration affect the neuron You must have the EC for K maintained at certain levels This is done by astrocytes which try to maintain the EC Whenever the concentration is high astrocytes kick in and cause depolarization The blood brain barrier makes it so that potassium cannot get into the brain A system that protects our brain and our CNS Lethal Injection Refer to ppt and the book The injection is injected into the blood stream and due to the blood brain barrier the injection travels to the heart where it stops the heartbeat and the blood supply to the brain Shaker K Channels What are the ion channels Which plays important role in the RMP It s just called resting ion channel and are open all the time and are primarily permeable to potassium ions That is due to the defect in one type of potassium ion channel Called shalter potassium ion channel Refer to structure from the ppt BIL 268 01222015 Lecture 4 Action Potential The purpose of the Nernst equation is to calculate the equilibrium potential Assumption is that the membrane is only permeable to that kind of ion You must have other types of ions in order to reach equilibrium Nernst equation is used because sometimes the membrane is permeable only to a specific kind of ion Along the different kinds of axons you can have axons without myein sheaths or axons that aren t covered by the gia ces I Properties of Action Potential Measurement of Action Potential There are two ways to record AP Intracellular recording tip of an electrode is stuck into the cell and a signal is recorded Extracellular recording the tip gets close to the extracellular surface of the neuron to detect the AP When you are recording the AP you need two electrodes Recording electrode and the reference ground electrode Action potential changes over time The amplifier is connected to the oscilloscope which shows the typical IC and EC recording What is the major difference between the records of the IC and the EC The shape of the recording as well as the amplitude of the recording The electrode tip in the cell generates huge AP ie microvolts vs miivots The recording is depending on how you connect the electrodes Why do we use the amplifier Because the electrodes have a very high resistance The recording would show a very small portion of the AP without being amplified The membrane potential increases until it reaches a certain threshold Once depolarization reaches the threshold it generates the action potential If the pulse isn t strong enough it causes the membrane potential to not reach the threshold There are two ways to define threshold You can use the electrical pulse and change it until you reach the threshold that generates an AP Or you can look at the membrane potential until you reach the membrane potential that triggers the AP 0 The lowest membrane potential that triggers the AP is the threshold There are different phases of the AP Rising Phase Falling Phase Overshooting Undershooting If you look at the entire AP you will notice that only the major portion is around 1 millisecond Generation of Action Potential You need to know what the threshold is and what the firing rate is The firing rate is the number of action potentials divided by the time in seconds The firing rate gives you the number of spikes and the time You start the stimulation and cause the membrane potential to increase a bit till it reaches MP Increasing the stimulus causes the AP 0 Either you generate AP or no AP I There is no in between If you record another neuron it could have different amplitude from the recording of the first neuron because its physical properties could be different Increasing the amplitude of the stimulus gives you more action potentials see graph on slide 4 The amplitude of the AP from a given neuron is the same During action potential there are two periods Absolute when you have one action potential that is due to your first stimulation o If you want to use the second stimulus and that stimulus falls within the absolute period it will not generate a second action potential 0 During this time no additional AP can be generated I The ion channels are inactive Relative when you use a second stimulus the amplitude of the stimulus has to be larger than he first one Why do you need a stronger stimulus to create an additional AP during the relative penod To compensate for the undershoot ll Mechanisms of Action Potential Depolarization Repolarization and Hyperpolarization The reference level is the membrane potential Usually a negative number If the reference membrane moves towards the more positive direction then it is considered to be depolarization or increasing the membrane potential Hyperpolarization is when you the membrane potential becomes more nega ve Repolarization is when you move towards the reference level Depolarization occurs due to the influx of sodium ions repolarization occurs due to the efflux of potassium ions The intercellular matrix has a higher concentration of potassium ions The voltage clamp is used to manipulate the membrane potential at a certain level You can increase the membrane potential and hold it at that level to look at the ion movements and so forth You can use the two electrodes and then a clamp electrode a third electrode to hold the voltage at a specific voltage and then observe the ions A giant axon squid was used The key thing is the ionic movement current flow across the cell membrane In the baseline anything plotted downward shows the influx of current 0 Causes depolarization of the cell membrane TTX puffer fish was used to observe that only the outward current was observed and the inward current disappeared o TTX blocllts voltage gated sodium ion channels TEA mushroom cap was used to observe that the outward current disappeared and the inward current was there 0 TEA blocllts voltage gated potassium ion channels I Not the same as the RM P How is the action potential explained using the current flow Conductance was measured The rising phase was due to the influx of sodium and the falling phase was due to the efflux of potassium VoltageGated Ion Channels The patch clamp First the electrode is made via a glass capillary which is heated and pulled to make the tip of it very small 15 microns in diameter The tip has to be polished Under the microscope a manipulator is used to gradually move the electrode under the microscope and the tip hits the outer surface of the membrane An electric current is passed through the tip and the resistance shows up on the screen during the pulse When you hit the membrane with the tip you dramatically increase the resistance and that current a very low current dramatically increases the voltage which goes very high 0 Indicator of the seal The manipulator can be used to move the electrode away and to tear a piece of the cell membrane 0 After the preparation the rest of the experiment can be conducted Patch clamp is different from voltage clamp and is more difficult For the voltage gated ion channel you will find four subunits and each subunit will have 6 alpha helixes with the pore loop The pore loop only allows the sodium ions to pass by The voltage is gated because the ion channel has a voltage sensor When changing the cell membrane potential we cause structural changes of the membrane proteins 0 When it is depolarized the voltage gated ion channels are opened When there is an influx of ions in the cell most of them are hydrated The potassium is unable to go through the ion channel because of it s complex and because it is too large for the ion channel The voltage gate opens during depolarization There are three important properties of voltage gated ion channels They open with little delay They stay open for about 1 ms a very short period of time before closing The channel can only be reopened once the membrane potential drops to the threshold TTX is a sodium voltage gated ion channel blocker Brevetoxin 3 is a sodium ion channel activator and keeps the channel open all the times Will cause in exhaustion and loss of function The voltage gated potassium channel also has its functions They have a similar structure to the sodium channels The channels open with a 1ms delay 0 Once the sodium ion channel has opened the potassium ion channel will open I One is influx and the other is efflux TEA is the potassium channel blocker Ionic Base of Action Potential You need to know what is the threshold and what the different phases are The sodium channels are inactive during the falling phase Overshoot is the same as the rising and the falling phases Undershooting is when the potassium channels open and all the sodium channels close Undershoot is below the RM P It s below because potassium is the major player and the movement of the potassium is towards the EP The driving force is the membrane potential minus the equilibrium potential for potassium Ill Action Potential Conduction Action Potential Conduction Along the Unmyelinated Axon Refer to the ppt for the diagram If a strong enough stimulation is given you can record the action potential Why do we never see the AP travelling back towards the cell body The AP falls in the reflective period which means it can t fall back The axon hillocllt is the spike initiation zone Action Potential Conduction Along the Myelinated Axon The axon is covered and due to the node of Ranvier the membrane has a very high concentration of voltage gated ion channels The action potentials can be generated and will jump to the next node The AP is very fast Saltatory Conduction Factors Influencing Conduction Velocity The physical characteristics of the axon affect the velocity The diameter and the myelin sheath affects the internal resistance Increasing the IR the AP conduction is reduced Spike Initiation Zones The axon hillocllt is the location of the spike initiation zone The zone depends on whether it s a typical neuron or a sensory neuron BIL 268 01272015 Lecture 5 Synaptic Transmission Are majority of the synapses electrical or chemical Chemical Electrical synapses respond faster in comparison to chemical synapses Everything related to escape behavior is electrical synapse Charles Sherrington coined the term synapse and received a noble prize for his work I Types of Synapses Electrical Synapse Gap Junction The two membranes are very close together and the top one is the presynaptic membrane and the bottom one is the post synaptic membrane A membrane protein is added connexon and is found The action potential comes down through the membrane resulting in depolarization and will accumulate in the ions and cross through the synapses Another name for the electrical synapse is gap junction Chemical Synapse Synaptic Structure You need to know the structure At the terminal of the axon you can see an enlarged area This is where you see the synaptic vesicles The vesicles are fused to form an active zone and release neurotransmitters Once the neurotransmitter gets in the cleft it binds to the extracellular side resulting in the change at the post synaptic neuron If you look at the presynaptic membrane it is thicker The postsynaptic membrane is also thick and is also known as the postsynaptic density Types of CNS synapses This is dependent on the location of the synapses Axon dendritic synapse axon somatic synapse axo axonic synapse Each axon is divided into different branches The post synaptic neuron receives multiple axons Gray39s Type I are asymmetrical and excitatory Excitation is depolarization Gray39s Type II are symmetrical and inhibitory Inhibitory is repolarization Neuromuscular Junction The presynaptic neuron is a neuron and the postsynaptic cell is a muscle cell and no a neuron If something wrong happens there is a risk of paralysis There is a large area is the motor neuron part ACh is the first neurotransmitter discovered ll Synaptic Transmission 3 types of neurotransmitters Don t worry about the structures You need to know the classifications Amino Acids Amines Peptides Neurotransmitter Synthesis and Storage Where is the location of the protein synthesis is in the cell body which is where the Rough ER is located and in the spiny dendrites No protein synthesis in the axon Amino acid and amines are found in the axon Neurotransmitter Release The action potential travels along the terminal and causes depolarization reflux of sodium The depolarization opens up the voltage gated Ca channel This will result in an influx of calcium Ca is high on the extracellular side and will trigger vesicle docllting Synaptic vesicle moving towards the membrane and will then release NT via exocytosis There are two types of snare proteins T snare proteins Target because one end is anchored in the membrane 0 The other end is floating around Neurotransmitter Receptors There are three types of NT receptors One is called NT gated ion channels 0 When the NT release binds to the EC side the ion channels are opened and move across the ion channel The NT do NOT move through the ion channel 0 The effects on the Post synaptic neuron can be of two ways I One response is called Excitatory Post Synaptic Potential EPSP and the other is called Inhibitory Post Synaptic Potential IPSP U The response depends on the NT or the NT receptors I What is the EPSP D You will see depolarization U The ACh binds to the receptors in the skeletal muscle Not always results in EPSP It s really depending on what is the membrane potential level at the time o If the MP lt 0 mV you get EPSP o If the MP gt 0 mV you get IPSP That means that for ACh If the MP 0 it will cause reversal potential 0 This is due to whether or not there is influx of sodium and efflux of potassium U Glutamate can only result in EPSP When glu binds to its receptors it causes an influx of sodium which causes EPSP I In IPSP there is a glycine NT channel U Release of glycine causes binding to the receptors which causes the influx of chloride ions U The negatively charged ions get in and cause hyperpolarization The second type is called G protein coupled receptors G PCR o The receptor is coupled with the G proteins intercellular side 0 The NT receptor is not an ion channel 0 The response is a bit slow because first it has to bind to the receptors and then affect the g proteins but it can amplify the response and be long lasting The third type is called auto receptors AR 0 The NT receptor is in the presynaptic membrane 0 Why I To stop the signal U Regulation I Too much glu can cause over excitation and can cause problems D It will kill the Post synaptic neuron and result in neuronal death in the brain Neurons in the brain cannot regenerate B One way is to try and control the NT release Release leads to bind in the membrane and causes additional release Neurotransmitter recovery and degeneration Release of the NT in the synaptic cleft Not all the NT binds to receptors because it moves via diffusion 0 Some llteeps fused away Reuptallte is when some of the NT re enters the presynaptic axon terminal Enzymatic destruction is inside the terminal cytosol or synaptic cleft and leaves ACh to inactive state Desensitization is when the binding between the NT and the receptor is poor Neuropharmacology The receptor antagonist which means that the NT blocks the receptors Receptor agonist which means that the NT binds to the receptors and mimic NT response Ill Synaptic Integration Dendritic Properties Cable lillte dendrites with a constant length What is the length constant o It basically means that the distance away from the original stimulus sites I The distance is measured to have a voltage of about 37 of the voltage recorded right at the stimulus site I When the voltage drops to that 37 it s called the Nernst constant I Equation is in the ppt n excitable dendrites you can find some voltage gated ion channels but not enough If you have one neuron that is 5 microns and another that is 10 microns then based on the given information you can determine that one neuron is smaller than the other Which neuron is more excitable o The second one I The larger one will have a lower internal resistance EPSP If the stimulus pulses are close together each pulse creates one EPSP and they get added up together NOT action potential Each AP generates one EPSP For a given NT there is mini EPSP with a fixed height If you have an EPSP with greater amplitude can you estimate how many SV are released 0 Yes you can I This is the quantal analysis which is used to determine a number of vesicles that are released during neurotransmission Shunting Inhibition What is it It s basically when the inhibitor synapse acts like a control If the synapse is not active the EPSP will pass through and reach above threshold and create Action Potential What happens if the synapse gets activated o It will be below threshold You can also look at the mechanism and explain it in three different ways Influx of chlorine ions 0 Which get in and cancel out the EPSP Length constant reduction will reduce the membrane potential Modulation They will modulate the effectiveness of ion channels Example is protein kinase binding to the potassium channel making it excitable enough to increase the membrane resistance BIL 268 01292015 Lecture 6 Neurotransmitter Systems On what criteria would you call a chemical a NT There are three criteria that you need to follow Need to learn NT synthesis and metabolism I How to identify NT Three Criteria A NT must be synthesized within a neuron and it must be stored within the neuron So in what way can you verify that this chemical is inside a NT The second way is that you can use immunocytochemistry and in situ hybridization For immunochemistry you can look at the protein level because it is related to the function If you can identify protein you can predict the function For in situ you have DNA and genetic code transcribed into the mRNA and from mRNA you have a protein Instead of looking at the very end product the protein you are looking at the mRNA By labeling the mRNA you can show what cell the label is on There are always advantages and disadvantages The chemical is synthesized inside a cell neuron and stored inside the neuron This chemical must be released by the neuron You can collect the NT and analyze it The third step is that the chemical will generate a response at the post synapse neuron Immunocytochemistry is localization of molecules to cells The antibody labeling can localize certain molecules to the cell If you have something with a label you can put in an antibody solution which will permeate the molecule and obtain a response Antibody Immunochemistry is based on antibodies Basically each antibody is composed of one heavy chain and one pair of light chain They have one end that binds to the antigen and are called antigen binding sites The other ends can attach to the fluorescent marllter Antibody is called Immunoglobulin lg 0 Has five different criteria s because the constant area structure is different How do you make the antibody The way is to use the peptide which you can put with the carrier You inject into the animal host After a short wait the animal body will eventually generate antibodies against that agent You collect the blood and purify it to get the antibody 0 Using that you can tag it with dye and be ready to use it One antigen can have more than one binding site Antibodyprotein binding When you use antibody you don t use just one antibody In many cases you use two different kinds of antibodies 0 Primary and secondary antibody I The secondary antibody has the marker In this case you can get brighter signals instead ofjust using the primary antibody You use the primary antibody which will bind to the protein The second antibody will bind to the primary antibody This magnifies the signal because you have several secondary antibodies to bind to the primary antibody Fish Sensory Epithelium Epithelium are the sheaths of auditory receptor cells in fish ears Bending of the hair bundles activated the cell When you use antibody labeling you use a dye lillte Alexa 488 phalloidin antibody which binds to the F actin The Alexa 488 is the green marllter If you look at the image you ll see that all the hair bundles got marllted green Green Fluorescent Protein It is a luminescent protein from jellyfish They contain GFP Tg lsl1 GFP Transgenic fish expressing the GFP The motor neurons in the brain and the spinal cord expressed the GFP Innervation in the left and the right ear In situ hybridization Localized synthesis of protein or peptide to a cell to detect mRNA In this case you need to know the sequence and based on the sequence you can design the probe The probe is complimentary to the mRNA so that it can bind to the mRNA A marker can be attached to the probe Studying Synaptic Mimicry In order to find out whether the chemical will generate the response at the post synapse neuron you can use a microelectrode Technique known as microionophoresis 0 Pull the tip of a glass capillary and make it really small and fill it with a candidate chemical solution 0 Move the tip very close to the synapse and pump the chemicals out 0 Then put the electrode to record it from the post synapse neuron to see when you can detect membrane potential change o If that chemical has any charge you can pass current to pump it in and then use pressure to pump it out Receptors Rules A NT binds to its receptors ACh NT will only bind to ACh receptors For a given NT there are different receptor subtypes The different receptor subtypes can have totally different functions One can be excitation and one can be inhibition Henry Dale came up with the Dale s Principle Proposed that for a given neuron it only contains one type of NT 0 Incorrect A given neuron can have a glutamate NT but can also have different types of NT Receptor Subtypes You need to know about ACh in great detail For the ACh there are two types Name them via agonist binds to the NT receptors and will generate very similar response and antagonist the blocker is going to take the position of the NT and block the response Nicotinic is the agonist and muscarinic is the agonist of the ACh receptor Glutamate has AMPA NMDA and Kainate The first two need to be known GABA A is ion channels and GABA B is G protein coupled receptors ACh has nicotinic receptors can be found in the membrane of skeletal muscle cells with Nicotine agonist and curare a blocker antagonist The binding of ACh to the receptors will open up the cation channels which will result in the influx of sodium and the efflux of potassium Depolarization of the skeletal muscle fibers will depend on the membrane potential which will cause muscle contraction Muscarinic receptors are muscarine agonists and atropine antagonist The binding of ACh to the muscarinic receptor will lead to the activation of G protein which will open the G protein gated potassium channel which will cause an efflux of potassium This will cause hyperpolarization of the cardiac muscle fibers which will lead to heartbeat reduction ll NT Chemistry Synthesis and Metabolism For ACh the neurons containing them are called cholinergic neurons Neurons that contain DA NE and E are called catecholaminergic neurons Serotonergic neurons contain 5 HT Amino acidergic neurons contain Glu Sly and GABA Some neurons contain ATP NO and CO The ATP acts as a NT and has its own receptors NO is a gaseous NT and it breaks down very fast Cholinergic Neurons Know everything and know the chemicals to synthesize ACh Know about ChAT Know how to break down ACh using ACh E Membrane and vesicular transporters The cell gets the chemicals in the EC environment Choline is transported into the cell via the choline transporter This transports against the concentration gradient and it needs energy and uses the concentration gradient for the sodium ions IC choline and acetyl CoA with ChAT are used to synthesize ACh Has to be packed into the vesicles The ACh gets accumulated in the vesicles and the NT is released 0 The enzyme breaks down the excessive NT and the whole process repeats itself Some of the ACh binds to the receptors Catecholaminergic Neurons They are involved in movement mood attention and visceral function Tyrosine is used to synthesize the three different types of NT DA NE E Don t worry about the structures they have the similar structures Know the name of the NT and the precursor molecules that are used to synthesize these molecules Serotonergic 5HT Neurons They39re amine NT They re derived from tryptophan They regulate mood emotional behavior and sleep Selective serotonin reuptake inhibitors 0 SSRIs Antidepressants I You have more serotonin excitation I Not having enough will lead to inhibition It is also related to the sleep cycle Ill Transmitter Gated Channels Basic Structure Pentamer There are five subunits for this ion channel You can find ACh binding sites in the alpha sub units Amino Gated Ion Channels Glutamate gated ion channels will generate EPSP Will be an excitation and depolarization There are two types 0 AMPA o NMDA So overall binding of the glutamate to the ion channels will open up the cation channels and will result in positive charged ions moving across the ion channels The difference is that for AMPA the ion channels only allow sodium ions and potassium ions NMDA allows sodium potassium and calcium Calcium eventually becomes the driving force Influx of calcium Binding of the glutamate to the NMDA receptor will not open the ion channel Because the magnesium ions are blocking the ion channel Binding has to be simultaneous with depolarization because it will release the magnesium ion and open up the ion channel It is voltage gated Where does the depolarization factor in Maybe there are AM PA receptors nearby Maybe the glutamate binds to AMPA and causes depolarization while binding to the NMDA receptors Amino acidgated ion channels The modulator is not a NT and is not an agonist or an antagonist Binding to the receptors modulates ion channel response They have their own binding sites Opening of ion channels is dynamic IV Gproteincoupled receptors and effectors Basic Structure The structure is a single polypeptide with seven membrane spanning alpha helices Basic mode of operation One is short cut and the second is the second messenger cascade The Shortcut Pathway The binding of the NT will affect the ion channel s fast and local Second Messenger Cascade The alpha binds to the enzyme and it affects many steps before finally reaching the ion channel It is slower but has an amplified response Phosphorylation and dephosphorylation You have two proteins that attach the phosphate group to the protein and remove the phosphate group from the protein Due to the attachment of the phosphate group to the original protein the structure will be changed Divergence and Convergence Each factor is kind of like the membrane proteins You start with a NT which can have different receptors Each receptor leads to different effectors 0 From one to many Or you can have different types of NT each binding to its own receptors and eventually affecting the same effector 2315 758 AM


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