Effects of 5α-cholestan-3-one on the synaptic vesicle cycle at the mouse neuromuscular junction.
We have investigated the effects of 5α-cholesten-3-one (5Ch3, 200 nM) on synaptic transmission in mouse diaphragm. 5Ch3 had no impact on the amplitude or frequency of miniature endplate currents (MEPCs, spontaneous secretion), but decreased the amplitude of EPCs (evoked secretion) triggered by single action potentials. Treatment with 5Ch3 increased the depression of EPC amplitude and slowed the unloading of the dye FM1-43 from synaptic vesicles (exocytosis rate) during high-frequency stimulation. The estimated recycling time of vesicles did not change, suggesting that the decline of synaptic efficiency was due to the reduction in the size of the population of vesicles involved in release. The effects of 5Ch3 on synaptic transmission may be related to changes in the phase properties of the membrane. We have found that 5Ch3 reduces the staining of synaptic regions with the B-subunit of cholera toxin (a marker of lipid rafts) and increases the fluorescence of 22-NBD-cholesterol, indicating a phase change within the membrane. Manipulations of membrane cholesterol (saturation or depletion) strongly reduced the influence of 5Ch3 on both FM1-43 dye unloading and staining with the B-subunit of cholera toxin. Thus, 5Ch3 reduces the number of vesicles which are actively recruited during synaptic transmission and alters membrane properties. These effects of 5Ch3 depend on membrane cholesterol.
Department of Normal Physiology, Kazan State Medical University, Kazan, 420012, Russia.
M R
MR
Kasimov
Department of Normal Physiology, Kazan State Medical University, Kazan, 420012, Russia.
A R
AR
Giniatullin
Department of Normal Physiology, Kazan State Medical University, Kazan, 420012, Russia.
A L
AL
Zefirov
Department of Normal Physiology, Kazan State Medical University, Kazan, 420012, Russia. Electronic address: fysio@rambler.ru.
A M
AM
Petrov
D000818:Animals; D002462:Cell Membrane / Q000187:drug effects* / Q000378:metabolism; D002776:Cholestanes / Q000494:pharmacology*; D002784:Cholesterol / Q000378:metabolism; D003964:Diaphragm / Q000294:innervation*; D004558:Electric Stimulation; D005089:Exocytosis / Q000187:drug effects; D005456:Fluorescent Dyes / Q000378:metabolism; D066298:In Vitro Techniques; D021962:Membrane Microdomains / Q000187:drug effects / Q000378:metabolism; D051379:Mice; D055369:Miniature Postsynaptic Potentials; D009469:Neuromuscular Junction / Q000187:drug effects* / Q000378:metabolism; D010791:Phrenic Nerve / Q000187:drug effects* / Q000378:metabolism; D017729:Presynaptic Terminals / Q000187:drug effects* / Q000378:metabolism; D011726:Pyridinium Compounds / Q000378:metabolism; D000644:Quaternary Ammonium Compounds / Q000378:metabolism; D009435:Synaptic Transmission / Q000187:drug effects*; D013572:Synaptic Vesicles / Q000187:drug effects* / Q000378:metabolism; D013997:Time Factors
D002776:Cholestanes; C073804:FM1 43; D005456:Fluorescent Dyes; D011726:Pyridinium Compounds; D000644:Quaternary Ammonium Compounds; D002784:Cholesterol; C007123:coprostanone
5α-cholestan-3-one; Cholesterol; Lipid raft; Neuromuscular junction; Neurotransmitter release; Synaptic vesicle cycle