Actin cytoskeleton depolymerization with clostridium spiroforme toxin enhances the secretory activity of rat melanotrophs.
1. We measured membrane capacitance (Cm) in cultured rat melanotrophs pretreated with Clostridium spiroforme toxin (CST), which specifically depolymerizes cortical filamentous actin (F-actin). Phalloidin staining confirmed that CST treatment depolymerised the F-actin. 2. In control cells, cytosol dialysis with 1 microM Ca2+i increased Cm by 23 +/- 4 % (n = 11) relative to the resting Cm 400 s after the start of patch rupture. In CST-treated cells the increase in Cm was 32 +/- 5 % (n = 15), not significantly different from controls. The rate of Cm increase was affected transiently by CST treatment, peaking at 1 min after patch rupture. The maximal rate of Cm increase was 4.27 +/- 0.85 fF s-1 (n = 12; measured 200 s after the start of patch rupture) in controls and 8.0 +/- 1.35 fF s-1 (n = 23; measured 75 s after the start of patch rupture) in CST-treated cells (P < 0.01). 3. In control cells cytosol dialysis with 0 microM Ca2+i decreased Cm by 9 +/- 3 % (n = 7), in CST-treated cells Cm increased by 11 +/- 3 % (n = 7) relative to resting Cm 400 s after the start of cytosol dialysis. The rate of change in Cm remained constant (controls: -1 to -2 fF s-1; CST treatment: 1-2 fF s-1). 4. Transient and sustained effects of CST treatment on changes in Cm at high or low [Ca2+]i, respectively, suggest a distinct role of cytoskeleton in Ca2+-dependent and Ca2+-independent changes in Cm. Transient enhancement of the rate of Cm by CST is consistent with a barrier role of cytoskeleton in regulated exocytosis. The sustained effect of CST on Ca2+-independent changes in Cm suggests cytoskeletal involvement in endocytosis.
Chowdhury
H H
HH
Laboratory of Neuroendocrinology-Molecular Cell Physiology, Institute of Pathophysiology, Medical School, p.p. 2211, 1001 Ljubljana, Slovenija.
Popoff
M R
MR
Zorec
R
R
D036002:ADP Ribose Transferases; D000199:Actins / Q000187:drug effects / Q000378:metabolism*; D000818:Animals; D001427:Bacterial Toxins / Q000494:pharmacology*; D002118:Calcium / Q000494:pharmacology; D003599:Cytoskeleton / Q000187:drug effects / Q000378:metabolism*; D004553:Electric Conductivity; D005089:Exocytosis / Q000187:drug effects / Q000502:physiology; D008297:Male; D008544:Melanocytes / Q000187:drug effects / Q000378:metabolism*; D008856:Microscopy, Fluorescence; D018408:Patch-Clamp Techniques; D010590:Phalloidine; D011108:Polymers / Q000378:metabolism; D051381:Rats; D017208:Rats, Wistar
D000199:Actins; D001427:Bacterial Toxins; D011108:Polymers; C054948:iota toxin, Clostridium spiroforme; D010590:Phalloidine; D036002:ADP Ribose Transferases; D002118:Calcium