• Synaptic vesicles of cholinergic neurons are loaded with the neurotransmitter acetylcholine ( ACh+ )

• The membrane of these synaptic vesicles contains an electrogenic H+ -ATPase that pumps H+ into the vesicle, together with ACh+ /H+ -exchangers ( vAChT ) and Cl– /H+ -exchangers ( CLC )

• The coupling stoichiometry for vAChT is one ACh + exchanged for 2 H+ , and for CLC is 2 Cl – exchanged for one H+ , with the H+ -ATPase hydrolyzing one ATP for 2 H + pumped.

• These vesicles exhibit a membrane potential Vsv = 80mV , inside positive , and a pH more acidic than cytoplasm by 1.5 units.

• Assume that the cytosol of the pre-synaptic terminal has the following properties :

  • [ ACh+ ]i = 1mM , [ H+ ]i = 0.1µM , and [ Cl– ]i = 10mM

• You predict that vesicular transporters fill synaptic vesicles with 2200 ACh + molecules

• You have calculated already how high the vesicular [ ACh+ ] can reach with the available electrochemical gradients , rev EAch

• The solubility limit for ACh+Cl –is ~100mM.

1. Determine the driving force for the exchanger ( vAChT ) when the inside of the synaptic vesicle reaches this solubility limit

2. Calculate how many ACh+ molecules will be stored inside the vesicle at this limit

   • let vesicle volume = $40\ast {10}^{-21}Liters$$40 * 10^{-21}\ Liters$

   $$\text{total moles}=\frac{100\ast {10}^{-3}moles}{Liter}\ast 40\ast {10}^{-21}Liter=4\ast {10}^{-21}moles$$
   $$4\ast {10}^{-21}moles\ast \frac{6.022\ast {10}^{-23}molecules}{mol}=2408.9molecules$$

3. Describe a likely outcome of ACh+ uptake in this vesicular system

   • the vAChT exchanger will stop running once the driving force for ACh to enter is zero

4. Calculate how many ACh+ molecules will be stored inside the vesicle then

   • 2408.9