Intro to Neurophysiology

Introduction to Neurophysiology

Recap of Previous Class

  • Focus was on G Protein-Coupled Receptors (GPCRs), which are transmembrane receptor proteins coupled with G proteins.

  • Three main types of G proteins:

    • G alpha (main driver of GPCR signaling)

    • G beta

    • G gamma

  • Four major G alpha families:

    • G alpha s: Stimulatory; increases cyclic AMP (cAMP) production by activating adenylate cyclase.

    • G alpha i: Inhibitory; decreases cAMP production by inhibiting adenylate cyclase.

    • G alpha q/13: Modulatory; stimulates phospholipase to generate inositol triphosphate (IP3) and diacylglycerol (DAG), leading to increased intracellular calcium by opening calcium channels in the endoplasmic reticulum.

    • G alpha 12/13: Involved in cytoskeletal modulation and cellular locomotion by activating Rho family proteins (e.g., Rho and Rac).

  • Beta and gamma subunits operate alongside G alpha but have distinct roles, mainly involving ion channels and interferon kinase activation.

Termination of GPCR Signaling

  • GPCR signaling is terminated via phosphorylation by GPCR kinases (GRKs).

  • Phosphorylation leads to the recruitment of beta-arrestin, resulting in receptor desensitization or internalization.

  • The diversity of GPCRs enables neurotransmitters (e.g., glutamate) to have varied effects due to different receptor types (ionotropic vs metabotropic receptors).

  • Ligands can induce biased GPCR signaling, favoring either G protein activation or beta-arrestin pathway activation, which has drug development implications.

Upcoming Quiz

  • Topics to focus on for the quiz include:

    • Dendritic spines and their function in the CNS.

    • Key non-neuronal cells in the brain:

    • Astrocytes

    • Microglia (M1 inflammatory vs. M2 anti-inflammatory)

    • Oligodendrocytes

    • Schwann cells

    • Review cranial nerves and spinal cord levels.

  • The quiz is due on Saturday with a time limit of approximately 15 minutes.

  • Emphasized importance of completing the quiz in one sitting for better retention of material.

Lecture Objectives

  1. Explain how neuroscientists manipulate neuronal activity using chemogenetic and optogenetic approaches.

  2. Identify and describe proteins involved in neurotransmitter release at the synapse.

  3. Analyze the roles of specific ion channels during each phase of the action potential.

  4. Apply the Nernst equation to calculate equilibrium potential for individual ions.

Manipulating Neuronal Activity

Chemogenetics

  • DREADD receptors (Designer Receptors Exclusively Activated by Designer Drugs) are used for manipulation of neuron activity.

    • Introduces a GPCR activated by Clozapine N-Oxide (CNO), which itself has no effect on other receptors.

    • Types of DREADD receptors:

    • hM3 Dq (stimulatory, coupled with G alpha s): Excites neurons.

    • hM4 Di (inhibitory, coupled with G alpha i): Inhibits neuronal activity.

    • Potential to genetically manipulate animals to express DREADD receptors selectively in certain neuron populations.

    • Specific promoters (e.g., SCL1 for adult-born neurons) can drive expression selectively.

  • Expression can be verified with fluorescent proteins for better targeting.

  • Allows for the study of neuronal function in behaviors by controlling activity through CNO administration.

Optogenetics

  • Introduces light-sensitive ion channels:

    • Channelrhodopsin: Opens sodium channels in response to blue light (excitatory).

    • Halorhodopsin: Opens chloride channels in response to blue light (inhibitory).

  • Enables real-time modulation of neuronal activity, great for observing behavior changes immediately in response to neural activation or inhibition.

  • Optogenetics operates with distinct light wavelengths for excitation and inhibition, overcoming some limitations of DREADDs.

Neurotransmitter Release Process

  • Action potential triggers calcium channels at axon terminals, causing synaptic vesicles to fuse with the presynaptic membrane and release neurotransmitters via exocytosis.

  • SNARE proteins are crucial in vesicle docking and fusion during neurotransmitter release.

    • Key SNARE protein: Synaptotagmin, acts as a calcium sensor that facilitates vesicle fusion upon calcium influx.

  • Termination of the signaling includes neurotransmitter breakdown or reuptake through transporter proteins.

  • Autoreceptors on presynaptic membranes provide negative feedback by monitoring neurotransmitter levels to regulate release.

Phases of Action Potential

  1. Resting potential: Established by potassium leak channels and sodium-potassium pump maintaining ion gradients.

  2. Depolarization: Triggered by ligand-gated sodium channels, which allow sodium to enter.

  3. Action potential threshold: When enough sodium channels open, voltage-gated sodium channels fully activate.

  4. Repolarization: Involves the opening of potassium channels resulting in potassium efflux, caused by sodium-potassium pump restoration.

  5. Hyperpolarization: Caused by prolonged opening of potassium channels then returns to resting potential.

Nernst Equation

  • Used to calculate equilibrium potential () determining the voltage at which there is no net ion movement: Eion=RTZFln⁡([Ion]out[Ion]in)E{ion} = \frac{RT}{ZF} \ln \left( \frac{[Ion]{out}}{[Ion]_{in}} \right)

    • Where:

    • Eion_{ion} = Equilibrium potential for ion

    • R = Ideal gas constant 8.314 J/(mol K)

    • T = Temperature in Kelvin

    • F = Faraday's constant 96485 C/mol

    • Z = Charge of the ion

  • Example Calculation:

    • For calcium, if at rest, equilibrium is 35.42 mV (calculated) using concentrations, R, and F as discussed in lecture.

Summary of Key Concepts

  • Manipulation of neuronal activity using chemogenetic (DREADDs) and optogenetic techniques.

  • Understanding neurotransmitter release mechanisms through SNARE proteins.

  • Mastery of action potential phases and the Nernst equation for determining equilibrium potentials.