18.Presynaptic mechanisms

Neurophysiology XVIII: Presynaptic Mechanisms

18.1 Overview of Fast Synaptic Transmission

  • Process of Fast Synaptic Transmission:

    1. Action potential invades presynaptic terminal.

    2. Voltage-gated Ca2+ channels open.

    3. Influx of Ca2+ triggers the fusion of vesicles with the plasma membrane.

    4. Neurotransmitter is released into the synaptic cleft.

    5. Neurotransmitter opens postsynaptic ligand.

18.2 Model Systems for Studying Synaptic Transmission

  • Studying Areas:

    • Central nervous system

    • Peripheral nervous system

  • Key Experimental Preparations:

    • Neuromuscular Junction: One of the first studied for transmitter release.

    • Calyx of Held Synapse: Found in the mammalian auditory system, resembles the calyx of a flower.

    • Squid Giant Synapse: Found in the stellate ganglion, responsible for triggering action potentials in the squid giant axon.

18.3 General Properties of Synaptic Transmission

  • Studying EPSPs:

    • Stimulating pyramidal neurons in the neocortex releases glutamate; each spike evokes EPSPs.

    • Amplitude of EPSPs varies across trials.

    • Average EPSPs show a characteristic fast rising phase and slower decaying phase.

    • Calyx Synapse Analysis:

      • Postsynaptic response measured during presynaptic spiking under voltage clamp.

      • Observe excitatory postsynaptic current (EPSC) and synaptic delay.

18.4 Calcium Dependence

  • Importance of Squid Giant Synapse:

    • Allows manipulation of the presynaptic terminal and measurement of transmitter release.

    • Experiments led to the hypothesis that Ca2+ is essential for transmitter release.

  • Evidence for Ca2+ Dependence in Synaptic Transmission:

    1. TTX Blockage: Transmission blocked by Na+ channel blocker, TTX, requiring action potentials.

    2. Depolarization in TTX: Allows release without Na+ influx.

    3. K+ Channel Blocker TEA: Depolarization still triggers release, showing K+ efflux is not required.

    4. Extracellular Ca2+ Removal: Inhibits transmitter release, underscoring the necessity of Ca2+.

  • Density of Calcium Current:

    • High concentrations of Ca2+ lead to increased fluorescence, representing influx.

18.5 Ca2+ Channels Linked to Transmitter Release

  • Squid Giant Synapse:

    • Rudolfo Llinas' Work (1989): Identified that transmitter release is due to P-type Ca2+ channels, blocked by funnel web spider toxin (FTX).

  • Mammalian Synapse Studies:

    • Cerebellar Synapse Observations:

      • Monitored presynaptic Ca2+ with furaptra dye.

      • N-type channel blockade reduced Ca2+ influx and synaptic transmission.

      • P-type channels significantly decreased transmission with complete inhibition.

  • Frog Neuromuscular Junction:

    • N-type Ca2+ channels trigger synaptic transmission.

    • Blockade of N-type channels with ω-conotoxin inhibits presynaptic Ca2+ increases.

18.6 Relationship Between Ca2+ and Release Rate

  • Model for Release Rate Calculation:

    • Protein X activates vesicle fusion by binding Ca2+.

    • Release rate is proportional to Ca2+ binding, leading to sigmoidal relationships at higher Ca2+ concentrations.

  • Historical Experiments:

    • Dodge and Rahamimoff (1961): Demonstrated cooperative Ca2+ release rate at the frog NMJ with a calculated cooperativity value of n=4.

    • Differences Identified in Squid Synapse: n=3, showcasing non-linear activation of Ca2+ currents affecting release.

18.7 Ca2+ Concentrations in Cells

  • Homeostasis of Ca2+:

    • Intracellular concentrations maintain low levels (20-200 nM).

    • Brief high concentrations appear near ion channels during influx, dispersing rapidly.

  • Mechanisms for Ca2+ Dispersal:

    • Rapid binding to proteins.

    • Transport into smooth ER or mitochondria

18.8 Release Mechanisms

  • Vesicle Transport & Recycling Overview:

    • Neurotransmitter transport into vesicles via H+-dependent anti-port system.

    • Vesicle docking, priming, and exocytosis processes outlined.

  • Key Proteins Associated with Release Mechanisms:

    • Vesicle-Associated Proteins:

      • Synapsin, Rab3, Synaptophysin, Synaptotagmin (Ca2+ sensor), Synaptobrevin (v-SNARE).

    • Cytoplasmic Release Proteins:

      • NSF, SNAPs, and Calcium channels.

  • SNARE Hypothesis:

    • Involves processes that promote vesicle fusion in response to Ca2+.