Integrative Cell and Tissue Biology: The NEURON

BIOM2011/3: Integrative Cell and Tissue Biology - The Neuron

Learning Objectives for Neuronal Physiology

• Structural Identification: Define and identify dendrites, axon, axon hillock, soma, synapses, and the synaptic cleft on a neuronal diagram.

• Neurotransmitter Criteria: Define the specific characteristics required for a chemical to be classified as a neurotransmitter.

• Chemical Neurotransmission Sequence: Describe the temporal sequence of events starting from the arrival of depolarization at the pre-synaptic membrane to the generation of a graded potential at the post-synaptic membrane.

• Signal Contrast: Contrast the generation and conduction of graded synaptic potentials versus action potentials, including their specific locations on the neuron.

• Ionic Basis of Synaptic Potentials: Describe the ionic mechanisms for inhibitory and excitatory post-synaptic potentials (IPSPs and EPSPs) and how they alter transmission.

• Transmission Comparison: Compare electrical and chemical synaptic transmission regarding velocity, fidelity, and the capacity for neuromodulation (facilitation/inhibition).

• Inhibition Types: Distinguish between postsynaptic and presynaptic inhibition with examples.

• Synaptic Plasticity: Contrast the characteristics of short-term versus long-term synaptic plasticity.

Historical Foundations and Functional Value

• Advantages of Neuronal Signaling:

  • Fast: Signal conduction ranges from $0.5-120\,m/s$.

  • Inexpensive: Requires only small amounts of signaling molecules.

  • Directional: Sent to specific targets over long distances.

  • Selective: Signals are small, discrete, and highly targeted.

• Variability: Neurons are the most variable cells in the body in terms of shape, size, responses, functions, and modifiability.

Neuronal Structure and Morphology

• The Dendrite:

  • Originates in the soma.

  • Displays simple to complex branching structures.

  • Variable in diameter and length.

  • Often covered in small protuberances called spines (dendritic spines).

  • Comprises the majority of the neuron's membrane area and receives the bulk of synaptic inputs.

• The Soma (Cell Body):

  • Contains the nucleus and metabolic machinery.

• The Axon Hillock:

  • The proximal end of the axon where the signal is initiated.

• The Axon:

  • An extension carrying signals to synapses.

  • Variable diameter and length (can be very long).

  • Generally has few or no branches until the terminal end (terminal branches).

• The Synapse:

  • Specialized axon endings for communicating signals between cells.

  • Presynaptic Terminal: Small swelling (~$0.5-1\,\mu m$ diameter) at the axon end.

  • Mitochondria: Present to support energy-dependent processes such as exocytosis.

  • Synaptic Vesicles: Small membrane-bound packets containing neurotransmitters.

  • Presynaptic Density (Active Zone): A collection of proteins for locating, exocytosis, and recycling vesicles.

  • Synaptic Cleft: A narrow extracellular gap (~$30-50\,nm$) between membranes.

  • Postsynaptic Density: A collection of proteins including receptors, scaffolding, and signal transduction molecules.

Membranous and Electrical Properties

• Ionic Barrier: The biplanar lipid structure of the cell membrane is non-permeable to charged ions. Movement depends on the ion's charge, chemical gradient, and electrical distribution.

• Membrane acts like a capacitor, storing charge and contributing to the electrical excitability of neurons, which is essential for the generation and propagation of action potentials.

• Ion Channels: Proteins with pores that allow selective ion passage based on charge and size.

• Electrical Definitions:

  • Current (I): Rate of movement of charged particles. Biological magnitude is ~$picoA$ ($10^{-12}\,A$).

  • Potential/Voltage (E or V): Force acting on particles due to charge imbalance (the "battery"). Biological magnitude is ~$milliV$ ($10^{-3}\,V$).

  • Conductance (g): Ease of movement. Biological magnitude is ~$nanoS$ ($10^{-9}\,S$).

  • Resistance (R or Omega): Inverse of conductance ($R = 1/g$). Biological magnitude is ~mega̦ ($10^{6}\,\Omega$).

  • Capacitance (C): Ability to store charge. Biological magnitude is ~$microF$ ($10^{-6}\,F$).

• Ohm’s Law: $I = gV$ or $V = IR$. If $g = 0$, no current flows regardless of voltage. If $V = 0$, no current flows regardless of conductance.

• Calculations Examples:

  • If a single $K^+$ channel has $g = 50\,picoS$ and a neuron has R = 100\,mega̦, the number of open channels is calculated as follows:

    • $R_{\text{channel}} = 1 / (50 \times 10^{-12}) = 2 \times 10^{10}\,\Omega$.

    • $Channels = (2 \times 10^{10}) / (100 \times 10^6) = 200\,channels$.

  • To cause a $+10\,milliV$ shift in a neuron with 100\,mega̦ resistance: $I = V/R = (10 \times 10^{-3}) / (100 \times 10^6) = 100 \times 10^{-12}\,A = 100\,picoA$.

Resting Membrane Potential (RMP)

• Definition: The RMP is the potential difference where the inside of the membrane is negative relative to the outside, typically ranging from $-90$ to $-45\,mV$.

• Constant flux of K+ Na+ and Cl- throught non-voltage dependent ion channels

• Ionic Equilibrium Potential ($E_{ion}$): The electrical potential that exactly counter-balances the concentration gradient, resulting in no net ionic flow. Calculated using the Nernst Equation.

• Typical Ion Distributions (at $37^{\circ}C$):

  • $K^+$: Outside: $5\,mM$; Inside: $100\,mM$; Ratio: $1:20$; $E_{K} = -80\,mV$ .

  • $Na^+$: Outside: $150\,mM$; Inside: $15\,mM$; Ratio: $10:1$; $E_{Na} = 62\,mV$.

  • $Cl^-$: Outside: $150\,mM$; Inside: $13\,mM$; Ratio: $11.5:1$; $E_{Cl} = -65\,mV$.

  • $Ca^{2+}$: Outside: $2\,mM$; Inside: $0.0002\,mM$; Ratio: $10,000:1$; $E_{Ca} = 123\,mV$.

• Maintenance of Gradients:

  • Na-K ATPase: Moves $3\,Na^+$ out and $2\,K^+$ in. Uses up to $70\%$ of brain ATP.

  • Ca pump: Moves $Ca^{2+}$ into internal stores or out of the cell using ATP.

  • Cl cotransporter pump: Can move Cl- in and out of the cell

  • Goldman Hodgkin Katz (GHK) Equation: Used to calculate RMP based on multiple ions and their permeabilities.

• Regulation: RMP is predominantly determined by $K^+$ due to high permeability. Extracellular $[K^o]$ is buffered by glial cells via gap junctions.

• Signaling Conventions:

  • Inward Current: $+ve$ ions in or $-ve$ ions out. Causes Depolarization in RMP.

  • Outward Current: $+ve$ ions out or $-ve$ ions in. Causes Hyperpolarization in RMP.

• Patch Clamping:

  • Technique used to measure the ionic currents across the membrane of cells. Salt solution filled glass recording electrode pushed onto the membrane creating a tight seal.

  • Whole Cell Recording: More suction makes membrane break in electrode tip, allowing low resistance electrical access to all ion channels on neuron surface.

  • Membrane patch recording: Small patch of membrane inside electrode tip is pulled away from neuron, only ion channels are recorded.

The Action Potential (AP)

• Initiation: The "decision" to fire occurs in the axon initial segment. Requires depolarization from resting (~$-60\,mV$) to threshold (~$-40\,mV$).

• Phases of the AP:

  • Rising Phase: Threshold triggers rapid opening of voltage-gated $Na^+$ channels. Positive feedback loop: $Na^+$ entry causes further depolarisation. Positive charge accumulates on inside of cell membrane, membrane potential become more postitive.

  • Overshoot: Membrane potential ($MP$) moves toward $E_{Na}$.

  • Peak: $Na^+$ channels inactivate; voltage-gated $K^+$ channels open slowly, causing $K^+$ efflux.

  • Falling Phase: gNa decreases, gK increases.

  • Undershoot (After hyperpolarization): All Na+ channels close and open K+ channels hyperpolarises the membrane potential below the RMP. $K^+$ conductance hyperpolarizes the $MP$ below RMP toward $E_{K}$.

• Channel Dynamics:

  • $Na^+$ channels activate for $1-2\,ms$ then inactivate until membrane hyperpolarises.

  • $K^+$ channels open slower and stay open longer.

  • Macroscopic current is the average of many asynchronous single-channel "all-or-none" openings.

• Initiation: AP travels from the axon initial segment (Na+ channel density high) down to the terminal and back into the soma/dendrites.

• Sensory afferent: In peripheral tissues, fires when stimulus reaches threshold for AP

• Intensity of sensation: Number of firing afferents x firing rate x effect duration

• Conduction Velocity Factors:

  • Axon Diameter: Wider diameter equals faster speed (e.g., squid giant axon).

  • Myelination: Produced by Schwann cells (PNS) or oligodendrocytes (CNS). Insulation prevents leakage and reduces capacitance.

  • Nodes of Ranvier: Breaks in myelin with high density of $Na^+$ and $K^+$ channels (stained with Caspr at paranodes). Enables saltatory conduction, which is jumping of nerve impulse.

• Pathology: Demyelinating diseases include Multiple Sclerosis and Acute Demyelinating Polyneuropathy.

• Pharmacology/Toxins:

  • Tetrodotoxin (TTX): From puffer fish, blocks voltage-gated $Na^+$ channels (puffer fish themselves have mutant, insensitive channels). Known as the ingredient in "zombie powder."

  • Local Anesthetics: Also block $Na^+$ channels to prevent AP generation.

Sensory Coding and Information

• Firing Rate: Information is conveyed via the frequency and number of APs (e.g., regulates muscle contraction strength or intensity of sensory stimuli).

• Receptive Fields: Each sensory afferent relays stimuli from a specific body area.

• Adaptation: Decrease in firing rate during continued stimulation.

  • Phasic Receptors: Rapidly adapting, strong response at stimulus onset.

  • Tonic Receptors: Slowly adapting, continuous response during stimulus.

Synaptic Transmission: Electrical and Chemical

• Synaptic transmission occurs at synapses which are specialised contacts between excitable cells.

• Chemical Synaptic Potentials (graded)

  • Synapses occurs on dendrites, soma, axon hillock, and other synapses.

  • Synaptic potential produced by synapses

  • SPs are small changes in membrane potential caused by ions moving through ion channels activated directly or indirectly by neurotransmitters.

• Electrical Synapses:

  • Connection via Gap Junctions (narrow $3\,nm$ gap) which contains connexons.

  • Connexons: Protein channels allowing direct ion/molecule flow. Large pores to allows direct passaging of ions and small molecules.

  • Features: Fast, bi-directional, fail-safe. Important in cardiac/smooth muscle and glial cells.

  • Modulation: High intracellular $Ca^{2+}$ closes connexons.

Neurotransmitter: Chemicals used to relay, amplify, and modulate electrical signals between neuron and another cell.

• Neurotransmitter Definition Criteria:

  1. Synthesised endogenously in the presynaptic neuron.

  2. Available in sufficient quantity to exert an effect.

  3. Mimics endogenous release when externally administered.

  4. Features a biochemical mechanism for inactivation.

• Neurotransmitter Reponses

  • Ligand-gated ion channels: Receptor proteins with integral ion pore, open rapidly when transmitter molecule binds to receptor site

  • G-protein coupled receptors: receptor protein without integral ion pore, activates separate G-proteins and then effector proteins such as G-protein gated ion channels.

• Neurotransmitter Classes:

  • Amino Acids: Glutamate (Glu - excitatory), GABA and Glycine (GABA,Gly - inhibitory). (3G)

  • Amines: Acetylcholine (ACh), Dopamine, Adrenaline, Noradrenaline, Serotonin (5-HT), Histamine. (Big 6)

  • Peptides: Cholecystokinin, Dynorphin, Enkephalins, Neuropeptide Y, Somatostatin, Substance P.

• Functions

  • Amino acids mediate fast excitatory and inhibitory synaptic transmission at most CNS synapses.

  • Ach mediates fast excitatory synaptic transmission at all neuromuscular junctions via nicotinic acetylcholine receptors.

• Synthesis and Storage:

  • Amines/Amino Acids: Synthesized in terminals; enzymes transported from soma. Subject to feedback inhibition and can be stimulated to increase activity through Ca2+ phosphorylation. Stored in small clear vesicles via pH-gradient-powered transporters.

  • Peptides: Synthesized in soma as propeptides; packaged in Golgi; transported via anterograde axonal transport in large dense-core vesicles. Made from precursor proteins in cell body, specific proteases cleave precursor into appropriate peptides.

• Release Mechanism:

  1. AP depolarizes presynaptic terminal from axon.

  2. Depolarisation causes voltage-gated $Ca^{2+}$ channels to open.

  3. $Ca^{2+}$ influx triggers exocytosis of neurotransmitter from vesicle, diffusion across synaptic cleft.

  4. Neurotransmitter binds to post synaptic receptor and opens ion channel, postsynaptic potential generated.

• Synaptic Vesicle Cycle

  • Exocytosis of synaptic vesicle content is for continous recycling of synaptic vescicles

  • Vesicle membrane endocytosed and refilled with transmitter

  • Filled vesicles docked near active zone

  • Docked vesicles primed for release through ATP dependent process

  • Ca2+ entering through closely located Ca2+ voltage-gated channels triggers fusion of synaptic vesicle membrane to presynaptic vesicle.

• AP evoked transmitter release is HIGHLY Ca2+ DEPENDENT

  • Blocked voltage-gated Ca2+ channels/removal of extracellular Ca2+ = no transmitter release.

  • Synaptotagmin: The $Ca^{2+}$ sensor in the vesicle membrane (exocytosis of neurotransmitter); may require 4 $Ca^{2+}$ molecules to bind.

• Quantal Theory: Transmission is based on unitary steps ("quanta"). A "miniature" postsynaptic potential represents the release of a single vesicle. More release = larger synaptic response

• Recovery and Degradation:

  • Post synaptic receptors desensitises if neurotransmitter present for too long

  • Diffusion, re-uptake into terminals or glia, or enzymatic breakdown (e.g., Acetylcholinesterase which is a target for nerve gases and pesticides).

Postsynaptic Potentials and Receptors

• Ionotropic Receptors (Ligand-Gated): Fast electrical responses after neurotransmitter binding.

  • For all amino acids, some amines like Ach on nicotinic acetylcholine receptor.

  • Excitatory, net effect is depolarisation (EPSP, driven by inward current ESPC): Receptors like glutamate (AMPA, NMDA), or Nicotinic ACh are permeable to $Na^+$ and $K^+$.

  • Inhibitory, net effect is hyperpolarisation (IPSP, driven by outward current IPSC): Receptors like $GABA_A$ and Glycine are permeable to $Cl^-$.

    • $GABA_A$ receptor subunits have many phosphorylation sites for binding drugs which modulate receptor activity.

    • Major target for sedative drugs that increases $GABA_A$ current (more Cl-, more inhibition)

• Glutamate Receptors:

  • AMPA: Fast rising/falling; primary excitatory current.

  • NMDA: Slower; $Ca^{2+}$ permeable; blocked by $Mg^{2+}$ at rest; requires depolarisation to open.

• Metabotropic Receptors (G-Protein Coupled):

  • Slow responses due to delayed activation (>50\,ms delay) and long time course.

  • Activate second messenger systems which can modulate metabolism or open G-protein gated channels (usually $K^+$).

  • Can directly open/close G-protein gated ion channels via channel protein phosphorylation

Integration and Plasticity

• Only 1 vesicle is release at a time, neurons need to summate many excitatory and inhibitory inputs to determine if AP threshold is reached

• Inhibitory synapses are more dense on the soma/base of large dendrites. Synaptic inhibition can act as a gatekeeper with hyperpolarisation and shunting inhibition of excitatory synaptic responses

• Integration and Summation:

  • Spatial Summation: Summing inputs from different locations.

  • Temporal Summation: Summing inputs arriving at different times.

  • Shunting Inhibition: Opening $Cl^-$ channels near the soma to "short-circuit" excitatory currents from distant dendrites.

• Inhibition Types:

  • Postsynaptic: Hyperpolarization or shunting to reduce excitability.

    • Hyperpolarisation: RMP more + than E_Cl-, membrane potential will hyperpolarise when Cl- channels open

    • Shunting inhibition: RMP at or close to E_Cl-, membrane potential will not change significantly

  • Presynaptic: Controls transmitter release at individual synapses

    • Axo-axonic contact reduces $Ca^{2+}$ influx or increases $K^+/Cl^-$ conductance in the second terminal, specifically controlling release at that single synapse.

    • Reduced transmitter release caused by activation of K+/Cl- channels that decreses excitability in 2nd presynaptic channel or reduced opening of Ca2+ channels in 2nd presynaptic terminal (reduced exocytosis)

• Short-Term Plasticity (STP): Transient changes in synaptic response from the same synapse

  • Facilitation: Increased response due to rapid firing, reaches threshold more quickly

  • Depression: Decreased response due to vesicle depletion or other factors, threshold reached less quickly.

• Long-Term Plasticity (LTP/LTD):

  • Hebbian Theory: "Cells that fire together, wire together."

  • Long-Term Potentiation (LTP): A persistant increase in synaptic responses induced by high frequency stimulation.

  • Long-Term Depression (LTD): A persistant decrease in synaptic responses induced by low frequency stimulation.

  • LTP Induction:

    • Transmitter at CA3 to A1 is glutamate, activates AMDA and NMDA receptors

    • MP needs to be sufficiently depolarised by spatial and temporal summation for NMDA to allow Ca2+ into the postsynaptic cell

    • Increase in postsynaptic Ca2+ enhance activity of protein kinases

    • Requires high-frequency tetanus ($100\,Hz$) causing spatial/temporal summation to remove the $Mg^{2+}$ block from NMDA receptors, allowing $Ca^{2+}$ influx.

  • LTP Maintenance

    • Increase in CA1 synaptic inputs due to:

      • Postsynaptic change in AMPA receptos by phosphorylation of existing receptors and recruitment of additional AMPA receptors

      • Presynaptic increase of neurotransmitter released by retrograde actions of intercellular messenger like nitric oxide.

    • Permanent changes in synaptic responses = requires activation of genes to produce proteins