Mediation and modulation of transmitter release ppt

Course Work and Assessment Reminder

  • Topic: The development and function of the nervous system and its firing modalities across time scales (milliseconds to hours).

  • Discussion Statement: Elaborate on two types of functional neuronal signaling—one fast and one slow—detailing the molecular determinants of these processes. Additionally, describe a modulatory mechanism for one type and include key experimental evidence.

  • Course Work Details:

    • Test Date: Monday, 3rd November 12-12:45 (In-class, 15-20 questions).

    • Format: Questions requiring single-word or single-sentence answers; simple line diagrams.

    • Completion Time: 30 minutes.

    • Workshop Submission Due: Monday, 5th January.

Synaptic Transmission

  • Focus: Mediation and modulation of neuronal excitation in the nervous system (2025).

  • Chemical Transmission: Major method of intercellular signaling in the nervous system.

Key Points About Synaptic Transmission

  • Transmitter Release Mechanisms:

    • Overview of physiological, cellular, and molecular processes initiating synaptic transmission.

    • Identification of intermediates involved in fast transmitter release and biochemical evidence supporting these mechanisms.

    • Discussion on how transmitter release can serve as a template for modulation of neuronal signaling.

G-Protein Coupled Receptors (GPCR)

  • Family Overview:

    • Description of GPCR function and its diversity. (Reviewed in Year 2 course).

    • Use specific GPCR examples to illustrate modulation of neuronal excitation and interaction with ionic channels or vesicle trafficking proteins.

References for Further Reading

  • Standard texts and detailed reviews:

    1. Purves, D., Augustine, G., Fitzpatrick, D., Katz, L., LaMantia, A.-S., McNamara, J., & Williams, M. (Neuroscience). Sinauer.

    2. Sudhof, T.C. (2004). "The synaptic vesicle cycle". Annual Review of Neuroscience, 27, 509-547.

    3. Kononenko, N., & Hauke, V. (2015). "Molecular mechanism of presynaptic membrane retrieval". Neuron, 85, 484-496.

    4. Sudhof, T.C. (2013). "Neurotransmitter release: the last millisecond in the life of a synaptic vesicle". Neuron, 80, 675-690.

    5. Strock, J., & Diverse-Pierluissi, M. (2004). "Ca2+ channels as integrators of G-protein mediated signaling in neurons". Molecular Pharmacology, 66, 1071-1076.

    6. Yim YY, Zurawski Z, Hamm H. (2018). "GPCR regulation of secretion". Pharmacological Therapy, 192, 124-140.

Intercellular Signaling in the Brain

  • Conceptualization of Synapses: Early 20th century ideas by Cajal indicating that synapses facilitate all forms of intercellular signaling.

  • Key Experiment (Loewi, 1920s): Demonstrated chemical neurotransmission using heart perfusion.

    • Result: Perfusate from stimulated heart 1 influences the activity of heart 2.

  • Katz (1950s): Established foundational events at the neuromuscular junction defining synaptic transmission processes.

Dynamics of Synaptic Signaling

Key Mechanisms Involved

  • Presynaptic Action Potential: Required for triggering calcium influx necessary for synaptic signaling.

  • Post-Synaptic Response: Responding to pre-synaptic stimulation, resulting in membrane depolarization from -60 mV to +50 mV upon action potential triggering.

  • Calcium Influence: Lowered extracellular calcium diminishes neurotransmitter release efficacy.

Secretion Coupling Mechanisms

  • Action Potentials and Calcium Influx: Indispensable for neurotransmitter secretion and synaptic transmission.

Quantal Release Mechanism

  • Definition: Secretion of neurotransmitters in small packets referred to as quanta.

  • Observed Activity: Even unstimulated pre-synapse shows background miniature postsynaptic potentials (mEPPs).

  • Experimental Findings: Evoked postsynaptic potentials are multiples of mEPPs.

Background Activity Illustration

  • Quantitative Measurement: Statistical analysis of mEPPs provides insight into quantal release phenomena.

Vesicle-Mediated Transmitter Release

Evidence Supporting Vesicle Hypothesis

  • Electron Micrographs: Visual evidence of vesicle-mediated neurotransmitter release presents it as a primary mechanism.

  • Biochemical Testing: Purification of synaptic vesicles containing specific neurotransmitters (e.g., acetylcholine, glutamate).

  • Membrane Dynamics: Measurements indicating membrane area increase during synaptic excitation.

  • Vesicle Fusion Capture: Rapid freezing methods reveal vesicles at the plasma membrane during stimulation.

Calcium-Triggered Vesicle Fusion

  • Process: Vertex of neurotransmitter release via exocytosis, necessitating Ca2+ presence.

Vesicle Supply Replenishment

  • Mechanism: Continuous synaptic vesicle supply via recycling.

  • Experimental Approach:

    1. Marker Incubation: Non-membrane permeable markers utilized to trace synaptic vesicle activity.

    2. Mass Exocytosis Stimulation: High levels of potassium induce vesicle release.

    3. Recovery Protocol: Stopping stimulation and using imaging techniques for tracking.

Synaptic Vesicle Retrieval Routes

Pathways for Vesicle Recycling

  • Kiss and Run Mechanism: Direct and rapid way of synaptic vesicle reclamation.

  • Clathrin-Mediated Endocytosis: Slower and involves complex membrane dynamics.

  • Bulk Endocytosis: Engaged under high stimulation conditions to pool vesicles.

Key Proteins in Vesicle Cycling

  • Dynamin and Clathrin: Critical in the processes of vesicle entrapment, internalization, and reformation.

  • Adaptor Proteins: Facilitate clathrin recruitment and oligomerization necessary for vesicle retrieval.

Priming and Fusion Mechanisms in Active Zones

Organizational Features

  • Proximity and Timing: Critical aspects determining successful vesicle docking and release.

  • Key Proteins:

    • Munc-13: Activates syntaxin for efficient vesicle fusion.

    • RIM: Organizes vesicle release apparatus.

    • Complexin: Modulates SNARE complex formation.

Structural Considerations

  • Active Zone Dynamics: Venues for proximity and interaction among docking proteins, ion channels, and vesicle apparatus, leading to neurotransmitter release.

Mechanisms of GPCR Modulation

  • Overview: GPCRs serve as major modulators of synaptic transmission.

    • Represent about 1% of the human genome with diverse roles ranging from sensory signaling to neurotransmitter modulation (e.g., glutamate, neuropeptides).

Modulation Examples

  • G-Protein Interaction: Impact on transmitter release via calcium channels and receptor interaction dynamics.

Calcium Channel Regulation

  • Types of Channels: N, P, and Q type channels are major players in synaptic transmitter release.

  • Regulatory Mechanisms: Direct and indirect GPCR regulation leading to diverse pathways impacting neurotransmitter release.

Experimental Investigations of GPCRs

  • Experimental Models: Detailed analysis of GPCRs determining signal modulation, focusing on pathways to transmitter release.

  • Serotonin's Role: This neurotransmitter's facilitation highlighted in both presynaptic neurotransmitter release and postsynaptic potential dynamics.

Key Studies and Findings

  • Functional Pathways:

    1. Serotonin activates GPCR leading to inward current through K+ channel inhibition.

    2. cAMP-cascades leading to kinase activation and phosphorylation events, resulting in decreased potassium efflux and increased presynaptic calcium influx.

  • ** contrasting Effects**: Understanding that some GPCRs can inhibit rather than facilitate neurotransmission, adding complexity to the modulation of synaptic signaling.

Conclusion

  • Complex Interactions at Synaptic Sites: The interplay among various proteins (e.g. Syntaxin, Munc18, SNARE complex) and ion channels, mediated by GPCRs alongside calcium dynamics, forms a sophisticated system for neurotransmitter release.

  • Modele of Release: Highlighting the role of GPCRs in synaptic modulation offers insights into potential therapeutic targets for various neural disorders.