Mechanism of alpha-latrotoxin action at nerve endings of neurohypophysis.
The neurotoxin alpha-latrotoxin elicits spontaneous exocytosis of neurotransmitter from neurons and peptide hormones from endocrine cells. While the mechanism of action is not fully understood, both Ca(2+)-dependent and Ca(2+)-independent pathways participate in the facilitation of release, with the relative contribution of the pathways differing among neuronal and endocrine cell types. Here, we investigate the actions of alpha-latrotoxin on neuroendocrine nerve endings that emanate from central nervous system neurons and, therefore, are unique in that they possess properties of central nerve endings and endocrine cells. Using intracellular [Ca(2+)] measurements both calcium-independent receptors for latrotoxin (CIRL or latrophilin) and neurexin 1 alpha receptors were found to be functionally present. Interaction of alpha-latrotoxin with these receptors stimulated secretion of vasopressin and oxytocin neuropeptide. The secretory response was entirely dependent upon toxin-mediated extracellular Ca(2+) influx, although alpha-latrotoxin also consistently triggered mobilization of Ca(2+) from an intracellular store. The mobilization of intracellular Ca(2+) relied on alpha-latrotoxin-mediated Na(+) influx and was blocked by the protonophore FCCP, thereby implicating mitochondria as the Ca(2+) store being mobilized. Using the whole cell recording configuration of the patch clamp, we report that alpha-latrotoxin interaction with the CIRL receptor on these nerve endings resulted in ionic pore formation, generating unitary inward current steps of 20 pA and a channel conductance of approximately 220 pS in Ca(2+)-free saline. Thus, alpha-latrotoxin stimulates Ca(2+)-dependent exocytosis in neurohypophysial nerve endings through receptor interaction and insertion of Ca(2+) permeable membrane pores. While alpha-latrotoxin mobilizes intracellular Ca(2+) stores the elevation in [Ca(2+)] reached is insufficient to trigger measurable exocytosis.
Hlubek
Michael
M
Department of Molecular and Integrative Physiology, 7807 Medical Sciences II Building, University of Michigan, Ann Arbor, MI 48109-0622, USA.
Tian
Dequan
D
Stuenkel
Edward L
EL
D000818:Animals; D002118:Calcium / Q000378:metabolism*; D002478:Cells, Cultured; D005089:Exocytosis / Q000187:drug effects / Q000502:physiology; D006023:Glycoproteins; D007424:Intracellular Fluid / Q000737:chemistry; D008297:Male; D008564:Membrane Potentials / Q000187:drug effects / Q000502:physiology; D008928:Mitochondria / Q000378:metabolism; D009411:Nerve Endings / Q000187:drug effects* / Q000378:metabolism; D009419:Nerve Tissue Proteins / Q000378:metabolism; D009479:Neuropeptides; D009489:Neurosecretion / Q000187:drug effects* / Q000502:physiology; D010121:Oxytocin / Q000378:metabolism; D018408:Patch-Clamp Techniques; D010904:Pituitary Gland, Posterior / Q000187:drug effects / Q000502:physiology; D051381:Rats; D017207:Rats, Sprague-Dawley; D018000:Receptors, Peptide / Q000378:metabolism*; D013111:Spider Venoms / Q000494:pharmacology*; D014667:Vasopressins / Q000378:metabolism
D006023:Glycoproteins; D009419:Nerve Tissue Proteins; D009479:Neuropeptides; D018000:Receptors, Peptide; D013111:Spider Venoms; C034800:alpha-latrotoxin receptor; C112162:neurexophilin; D014667:Vasopressins; D010121:Oxytocin; C020763:alpha-latrotoxin; D002118:Calcium