Functional coupling between 'R-type' Ca2+ channels and insulin secretion in the insulinoma cell line INS-1.
Among voltage-gated Ca2+ channels the non-dihydropyridine-sensitive alpha1E subunit is functionally less well characterized than the structurally related alpha1A (omega-agatoxin-IVA sensitive, P- /Q-type) and alpha1B (omega-conotoxin-GVIA sensitive, N-type) subunits. In the rat insulinoma cell line, INS-1, a tissue-specific splice variant of alpha1E (alpha1Ee) has been characterized at the mRNA and protein levels, suggesting that INS-1 cells are a suitable model for investigating the function of alpha1Ee. In alpha1E-transfected human embryonic kidney (HEK-293) cells the alpha1E-selective peptide antagonist SNX-482 (100 nM) reduces alpha1Ed- and alpha1Ee-induced Ba2+ inward currents in the absence and presence of the auxiliary subunits beta3 and alpha2delta-2 by more than 80%. The inhibition is fast and only partially reversible. No effect of SNX-482 was detected on the recombinant T-type Ca2+ channel subunits alpha1G, alpha1H, and alpha1I showing that the toxin from the venom of Hysterocrates gigas is useful as an alpha1E-selective antagonist. After blocking known components of Ca2+ channel inward current in INS-1 cells by 2 microM (+/-)-isradipine plus 0.5 microM omega-conotoxin-MVIIC, the remaining current is reduced by 100 nM SNX-482 from -12.4 +/- 1.2 pA/pF to -7.6 +/- 0.5 pA/pF (n = 9). Furthermore, in INS-1 cells, glucose- and KCl-induced insulin release are reduced by SNX-482 in a dose-dependent manner leading to the conclusion that alpha1E, in addition to L-type and non-L-type (alpha1A-mediated) Ca2+ currents, is involved in Ca2+ dependent insulin secretion of INS-1 cells.
Vajna
R
R
Institute of Neurophysiology, University of Cologne, Köln, Germany.
Klöckner
U
U
Pereverzev
A
A
Weiergräber
M
M
Chen
X
X
Miljanich
G
G
Klugbauer
N
N
Hescheler
J
J
Perez-Reyes
E
E
Schneider
T
T
D017398:Alternative Splicing; D000818:Animals; D001464:Barium / Q000378:metabolism; D002121:Calcium Channel Blockers / Q000494:pharmacology; D015220:Calcium Channels / Q000235:genetics / Q000502:physiology*; D020908:Calcium Channels, R-Type / Q000235:genetics / Q000502:physiology*; D027682:Cation Transport Proteins; D018929:Cell Culture Techniques / Q000379:methods; D002478:Cells, Cultured; D004553:Electric Conductivity; D005947:Glucose / Q000494:pharmacology; D006801:Humans; D007328:Insulin / Q000378:metabolism*; D000078790:Insulin Secretion; D007340:Insulinoma / Q000378:metabolism*; D007515:Islets of Langerhans / Q000187:drug effects / Q000378:metabolism*; D051379:Mice; D018408:Patch-Clamp Techniques; D011189:Potassium Chloride / Q000494:pharmacology; D020033:Protein Isoforms / Q000235:genetics / Q000378:metabolism / Q000502:physiology; D013111:Spider Venoms / Q000494:pharmacology; D014162:Transfection; D014407:Tumor Cells, Cultured
C423009:CACNA1E protein, human; C498916:Cacna1e protein, mouse; D002121:Calcium Channel Blockers; D015220:Calcium Channels; D020908:Calcium Channels, R-Type; D027682:Cation Transport Proteins; D007328:Insulin; D020033:Protein Isoforms; C115521:SNX 482; D013111:Spider Venoms; D001464:Barium; D011189:Potassium Chloride; D005947:Glucose