Molecular Neuroscience Lectures 2024 PDF

Molecular Neuroscience 2024 (3 lectures). Overall. How does molecular knowledge inform us about nervous system.. Introduce broad principles to illustrate how molecular mechanism contribute o nervous system function.. Introduce some of the important molecules and processes in the nervous ystem an...

Molecular Neuroscience 2024 (3 lectures). Overall. How does molecular knowledge inform us about nervous system.. Introduce broad principles to illustrate how molecular mechanism contribute o nervous system function.. Introduce some of the important molecules and processes in the nervous ystem and why they are important to how it works..Describe the basic elements of synaptic signalling in a cartoon form.. Detail some of the principle modes of excitatory and inhibitory synaptic signalling.. Give specific examples of the molecules that organize excitation and inhibition.. Introduce some clinical relevance to Molecular Neuroscience. References Purves D, Augustine GA, Fitzpatrick D, Hall W, LaMantia A-S, McNamara JO, Williams SM (2007) Neuroscience, 4th edition. Sinauer Associates: Sunderland, MA 5th Edition available. Alberts B, Johnson A, Lewis J, Raff M, Roberts K, Walter P, (2008) Molecular Biology of the cell, 5th edition. Garland Science. 5th Edition available. Neuroscience can make a biochemist into psychiatrist and visa versa: template for success. Molecules Molecular Neuroscience Sub-cellular Cellular Systems Behaviours Physiological Pathological  Brain Complexity made simple? 30,000 genes DNA 100, 000 proteins Transcription 86 billion neurons 10-100 trillion synapses RNA Operate responses and signals on micro-millisecond time scale. Translation Retain or forget information for 10 secs to 70 years. Non vital organ as you are brain dead Proteins and survive. Lack brain activity develop relatively normally. Underlies number of major diseases Protein Function Alzheimer’s, Autism, Depression and Epilepsy Some of the tool box for investigating and understanding molecular Neuroscience. Regulated by Nerve activity Promoter studies. DNA Gene expression Yes Identifying mouse mutants. Disease forming mutations in humans. Maturation Yes cDNAs, PCR and in-situ hybridization RNA Gene profiling (e.g. microarrays, RNAseq). Alternate splicing Yes Antibody staining (western blotting Proteins or immunocytochemistry). Mature e.g. protein subunits come together. Transported to Yes where needed. Protein Function Targeted to correct domain or sub-cellular location Protein turnover Regulated (ubiquitination) (e.g.phosphorylation)  Promoters controlling nerve function Glutamate Glutamic acid Decarboxylase (GAD). Neurotransmitter 1 Neurotransmitter 2 GABA Glutamate is a natural amino acid GABA is synthesized Glutamate excitatory amino acid GABA is an inhibitory amino (Enhancers and Repressors) GAD gene is recognized by transcription factors to ensure gene is selectively expressed in GABA neurons Variation in morphology with a molecular basis: cytoskeleton. Born as a non-neuronal cell mature and cells-develop processes (see Katrin Deinhardt’s lectures). Polarity of processes (tend to be ended) -axon giving the presynaptic nerve terminal -dendrite giving the postsynaptic nerve terminal axon Retain this shape basic polarity static view but actually they are highly dynamic. Cytoskeleton is the important molecular component. +end -end ab Alpha/beta dimer build hollow tube G-actin monomers make F-actin filaments.  Organization of the major cytoskeleton components Post-synaptic Terminal Actin cytoskeleton often - + - associated in a cortical + Network enriched in Dendrite - + + terminal regions - Cell Body - Microtubules orientated unidirectionally - in the axon compartment but Axon bi-directionally in the dendrite Make tracks for Transport. + Neurofilament stabilizes axons + Tau Microtubule associated Pre-synaptic Terminal Protein-2 Regulatory proteins  Transport mechanisms:kinesin paradigm Giant axon big distances. Exclude cytoplasm. Purify Motor activity Activity Kinesin Protein gel Kinesin and other motors move things around. Binds to Cargo or protein that needs to be transported Binds to Microtubules. Uses ATP activity. Walk along the microtubule. Move protein to extremities of the processes where the - + major signalling happens.  Electrical and chemical potentials make neurons excitable. Na+/K+ATPase pumps 3 sodiums out 2 potassiums into neurons. More sodium outside (142 mM) than inside (10mM). More potassium inside (140mM) than outside (4mM). Major negative ion chloride is higher outside (103mM) inside (4mM). Minor ion Calcium outside (2mM) very low inside (100 nM). Excite Rest Inhibit Positive + out +++++++++++++++++ tive ---------------------------- ga in Ne - + Processes with polarity (axon and dendrite). Come close to each-other, potential to communicate. Synapse Information Using electrical communication to allow intercellular flow communication Using chemical communication to allow intercellular communication Protein holes that open and close in response to voltage changes. Sodium Channels made up of one Potassium Channels made up of one protein sequence that contains 4 domains. protein sequence that contains 1 domain. Each domain has a voltage sensor Each domain has a voltage sensor and ¼ of the pore. and ¼ of the pore. Come together in a tetramer to make a functional channel. Threshold for activation Threshold for activation about -50 mV about 0 mv Threshold for inactivation Threshold for inactivation about 0 mv +50 mV Select for Na+ Select for K+ Example of a functional Na+ channel Selectivity Filter Two ways to bridging the synaptic gap: electrical and chemical synapses Electrical Synapse. Signal passed direct electrical flow between two cells. Chemical Synapse. Electrical signal converted to a chemical signal and then back into an electrical signal.  Principles of chemical transmission. Stimulation Pre-nerve Secretion Synapse Diffusion Post nerve Receptor activation Signal Termination Diffusion away and uptake from synapse. Chemical transmission: a tale of ions, ion binding proteins, ion channel proteins, membrane fusion proteins and neurotransmitter (NT) transport proteins. Stimulated neuron opens ion channels including those that allow Ca2+ into nerve terminal. Ca2+ is sensed. Recognized by a protein that binds Ca2+ and changes its conformation. (Synaptotagmin). Change in conformation allows proteins SNARE proteins to promote fusion via a vesicle/plasmamembrane protein complex. Vesicle fuses with the plasma membrane, NT released and diffuses into the synaptic cleft. Receptors bind NT and these proteins are ion channels. NT binding opens (or is gates) receptor channel, allows ions to flow and change distribution across membrane. Excite by depolarizing the membranes {positive signal}. Inhibit by hyperpolarizing the membrane {negative signal}. Chemical signal is terminated by diffusion away or reuptake from the synaptic cleft. It happens much quicker than I can say it but how quick?  Initiating transmitter release by opening ion channels Stimulate nerve-open ion channels Calcium Channels open. depolarize membrane positive ion (Stimulation opens influx. ion channel selectively Ca2+ permeable to Ca2+). Gate Closed Gate Open  Ca2+ is coupled to vesicle fusion Ca2+ is sensed. Recognized by a protein that binds Ca2+ and changes its conformation (Synaptotagmin). Vesicle (v) and target membrane (t) SNARE proteins act as fusion promoting proteins (Make a vesicle/plasmamembrane complex). Synaptic SNARES Synptobrevin on synaptic vesicle Syntaxin on the plasma membrane SNAP-25 on the plasma membrane + 2 Ca Synaptotagmin vesicle protein. Ca2+ bound synaptotagmin -Protein domains that bind Ca2+ promotes vesicle SNAREs and -Changes Conformation when bound Ca2+ plasmamembrane SNAREs -Allows vesicle to see the signal from Ca2+ to complex using complementary protein interaction domains (i.e. coil-coil domains) this promotes fusion.  Neurotransmitter release, diffusion and reception. Vesicle fuses with the plasma membrane NT released (exocytosed) into the synapse and diffuses into and across the cleft. Two sides are very close (actually almost touching) Receptors Bind transmitter they are ion channels.  Receptor activation and transmitter action termination. Chemical signal is terminated by diffusion away Refil empty vesicles via neurotransmitter vesicle membrane transporters Local Reuptake by Receptor opens (or is gated) allows membrane transporters ions to flow in and change the distribution of ions. Inhibit by - + Excite depolarizing hyperpolarizing the the membranes membrane {negative {positive ions Na+ influx}. ion Cl- influx}.  In the brain synaptic sites are segregated and for reasons of function. Major excitatory synapse are on the dendrites and use Glutamate as the transmitter Yes No Major inhibitory synapses Axo-dendritic are on cell body and use synapse GABA or glycine as the transmitter. Cross and not make synapse Axo-somatic synapse  They look different and are molecularly distinct. Glutamatergic are often Asymmetric GABA/Glycine synapses are symmetric Active zone release site Vesicle glutamate Inhibitory amino acid transporter vesicle transporter (IAAT) Glycine PM Glutamate PM transporter transporter Stored glutamate Stored glycine Glutamate receptor Glycine receptor Thick specialization Flux Na+/Ca2+ Thin specialization organizer proteins Flux Cl-Inhibit organizer proteins Excite synapse. synapse. and cytoskeleton and cytoskeleton  Key Features on excitatory and inhibitory receptors Glutamate receptors Glycine receptors (GABA recpetors) Four subunits to make ion channel Five Subunits to make ion channel Glutamate binding site on outside Glycine binding site on outside Cation Channel Anion Channel Bind to molecules like PSD-95 on inside Bind to Gephyrin on the inside postsynaptic cell. postsynaptic cell Cartoon of the receptor protein Molecular model of receptor protein Agonist binding sites Post-synaptic membrane Intracellular receptor Gephyrin PSD-95 Binding-organizing molecules Find the cell body axon,dendrite and synapse in a real neuron. Camera Lucida of Cortical pyramidal neuron.  Organizing molecules are multi-domain proteins and can help with segregation. Pre-nerve Tag Tag Glutamate receptor Glycine receptor Post-synaptic nerve PSD-95 Gephyrin e.g. Neurexins a family of presynaptic tags Neuroligin 1 postsynaptic glutamatergic tag Neurligin 2 postsynaptic glycinergic tag Simple model as a primer for an complex synaptic glue. Glutamate Synapse GABA or Glycine Synapse Selective use of molecules can define synapse. (e.g. neuroligin 1, 3 , 4; excitatory: neuroligin 2 inhibitory) Make multiplex protein complex that are adhesive and bring about signalling. A real impact of molecular neurobiology in understanding and treating disease Molecule Disease Reason Drug Target indication Glutamate Cognitive decline Major routes for Activators act against receptors brain decline. (excitation) communication Inhibitors act against over-excitation PSD-95 Stroke Support over Novel approach (organize activity of reduce function by excitation) glutamate preventing PSD-95 receptors binding to receptor Glycine receptors Hyperekplexia: Mutation prevent Receptor (inhibition) increased startle proper glycine regulators used in response. receptor function pain and muscle thus reduced relaxants. inhibition. Gephyrin (organize Stiffman’s Auto-anitbodies IV introduction of inhibition) syndrome. against molecule of IgG. inhibitory synapses Modulate inhibition. Neurexin. Autism. Mutations effect Might think about Neuroligin. Autism and synapse gene therapy. schizophrenia development Modulate activity.. Introduce broad principles to illustrate how molecular mechanism contribute o nervous system function.. Introduce some of the important molecules and processes in the nervous ystem and why they are important to how it works..Describe the basic elements of synaptic signalling in a cartoon form.. Detail some of the principle modes of excitatory and inhibitory synaptic signalling.. Give specific examples of the molecules that organize excitation and inhibition.. Introduce some clinical relevance to Molecular Neuroscience. Q> which of the following would not contribute Different compartments of a cartoon neuron to the cytoskeleton underpins gross neuronal shape. are listed below. Where would you find i. Microtubules; ii.Tau; iii.MAP-2 A. MAP-2 iv. Synaptotagmin B. Tau C. Microtubules D. Actin A C E. Synaptotagmin B D