Lecture 19: Synaptic Vesicle Release and NMJ Pharmacology

What are the steps for synthesis and recycling of vesicles?

1. vesicle and peptide NT precursors and enzymes are synthesized in the cell and released from the golgi

2. vesicles travel through the axon on microtubule tracks via fast axonal transport. Peptide NTs are already in some vesicles

3. a nonpeptide NT is synthesized in the nerve terminal and transported into a vesicle

Where are peptide NTs synthesized, loaded, and transported?

synthesized at cell body, loaded into vesicles, and transported down axon

Where are non-peptide NTs synthesized and loaded?

synthesized in presynaptic terminal, loaded into vesicles

What are dense-core secretory granules?

• contain pre-synthesized peptides or peptide precursor when leaving cell body

• 100-200nm diameter

• randomly distributed in axon terminal

What are clear synaptic vesicles?

• travel down axon with no NT inside

• 40-50nm diameter

• take up non-peptide NT synthesized locally in nerve terminal

• docked at active zones

How does vesicular uptake of non-peptide NTs work with the vacuolar-type H⁺ pump/ATPase?

• catalyzes inward movement of H⁺ into vesicle

• resulting pH and voltage gradients energize NT uptake via neurotransmitter transporters

What are NT transporters?

• specific for different NTs

  • vesicular GABA transporter (vGAT)

  • vesicular glutamate transporter (vGlut)

What does fusion and recycling of synaptic vesicles look with SNARE proteins?

• molecular machinery for membrane-membrane fusion

• includes synaptobrevin (vSNARE), syntaxin and SNAP-25 (tSNAREs)

What does fusion and recycling of synaptic vesicles look with Synaptotagmin?

• Ca²⁺ sensor for exocytosis

What does fusion and recycling of synaptic vesicles look with Rab3?

• GTP-binding protein

• regulates synaptic release via GTP binding/hydrolysis

What are the steps for vesicle fusion and recycling?

1. vesicle docks near membrane

2. SNARE complexes form (vSNARE + tSNAREs) to pull membranes together

3. entering Ca²⁺ binds to synaptotagmin

4. Ca²⁺-bound synaptotagmin binds to SNARE complex and catalyzes membrane fusion

5. synaptotagmin dissociates from SNARE complex as [Ca²⁺]_i levels fall

6. a-SNAP and NSF use ATP hydrolysis to disassemble SNAREs

7. vesicle membrane and synaptobrevin endocytosed and recycled

What are the steps for local recycling of synaptic vesicles?

1. budding (exocytosed from endosome)

2. docking

3. priming

4. fusion along with Ca²⁺ entrance (1msec)

5. budding (endocytosed to endosome (10-20sec))

overall this takes 1 minute

What is Myasthenia Gravis and what are the symptoms? What happens during the disease?

• autoimmune disease

• Ptosis (droopy eyelid), difficulty speaking and swallowing, extremity weakness

• antibodies attack nAChRs in skeletal muscle NMJs

What was shown in the MEPP pathology in an animal model of Myasthenia Gravis?

• in the control, the nerve AP and muscle AP are normal

• in the EAMG (experimental autoimmune myasthenia gravis), there was a normal nerve AP, but no muscle AP

• in the table, the frequency and amplitude seem to majorly decrease from control to chronic myasthenia gravis. The quantum content drastically increases in chronic myasthenia gravis.

What synapses are target for toxins and drugs?

cholinergic NMJ synapses

What is the action of Acetylcholinesterase and what is its functional state?

• results in immediate termination of ACh actions at NMJ

• AChE either soluble, or bound to cell membranes and/or extracellular matrix

• choline left over after ACh breakdown is taken back up into presynaptic nerve terminal

  • via Na⁺ -coupled uptake

How do AChE inhibitors work?

AChE inhibitors (cholinesterase inhibitors) → ACh remains in synaptic cleft longer, nAChRs continue to be bound by ACh and activated → slower depletion of ACh by diffusion away from junctional folds → increased EPP amplitude, prolonged duration

What is Botulinum Toxin and what is its action?

• Botulism - weakness and paralysis of skeletal muscle

• inhibition of cholinergic motor neuron endings

What is Tetanus toxin and what is its action?

• tetanus (lockjaw) - general increase in muscle tension and rigidity

• prevents synaptic transmission from spinal cord inhibitory neurons

  • loss of inhibition of spinal cord motor neurons → increases stimulation of skeletal muscle contraction

What is true of both Botulinum toxin and tetanus toxin?

they both cleave SNARE proteins required for synaptic vesicle exocytosis