A Role Of A-latrotoxin In Cell Secretion

α-latrotoxin, a potent neurotoxin from black widao spider venom, is selectively targeted against vertbrates. α-latrotoxin forms dimers in solution that may assemble further into tetramers or higher order oligomers in the presence of divalent cations There are three distinct families of cell-surface receptors, Neurexin,latrophilin and PTPδfor α-latrotoxin. α-latrotoxin cause massive release of neurotransmitters from nerve terminals and cell exocytosis by froming pore and single transdunction mediated by receptor after binding to specific receptors. α-latrotoxin can mobilize the calcium store in cytoplasm and act on exocytotic machinery directly by receptors mediatied. Elucidation of precisely how α-latrotoxin works is likely to provide major insight into how cell vesicle exocytosis and the release machineries of neurotransmitters are regulated. In this study, by using capacitance measurement, microfluorometric measurement of calcium, flash photolysis of caged Ca²⁺, carbon fiber electrode, we studied the regulation mechanism of α-latrotoxin in regulation of the kinetics of exocytosis from single rat pancreatic βcells. Main results of the study are as fellows. We show α-latrotoxin can form pore with a unitary conductance of 0.13 nS on βcell membrane and induce Ca²⁺ influx form extracellular. α-latrotoxin could not release Ca²⁺ from calcium store. Trivalent cations, La3+, block the toxin pores. We find α-latrotoxin lowers the Ca²⁺ threshold required for exocytosis without the RRP change employed the ramp [Ca²⁺]i stimulation generated by continuous UV illumination; Using combined techniques of membrane capacitance measurement and Ca²⁺ uncaging, we show that α-latrotoxin accelerates the fusion kinetics of the readily releasable pool of vesicles, and α-latrotoxin shifts the Ca²⁺-dependence toward lower Ca²⁺ concentrations. In clacium free solution, we also show α-latrotoxin induce quantal release using carbon fiber electrode. The mutant toxin α-LTXN4C is a powerfu tool in the α-latrotoxin research. Despite the fact that this mutant has the same affinity for the receptors as the wild type α-latrotoxin, it fails to incorporate into the membrane and form pores as the wild type toxin does. α-LTXN4C can mimic the action on βcell exocytosis of α-latrotoxin. Moreover, the role of α-latrotoxin and α-LTXN4C dependent on divalent cations (Ca²⁺ or Mg2+). The different Protein Kinase C Inhibitors blocked the effect of either α-latrotoxin or α-LTXN4C in lowering the Ca²⁺ threshold for exocytosis, suggesting that PKC activation is required for the receptor-mediated modulation of the secretory machinery by α-latrotoxin. Our article propose that α-Latrotoxin, by binding to the membrane receptors, sensitizes the fusion machinery to Ca²⁺ and, hence, may permit release at low [Ca²⁺]i level. This sensitization is mediated by activation of protein kinase C.