The intracellular loop between domains I and II of the B-type calcium channel confers aspects of G-protein sensitivity to the E-type calcium channel.
Neuronal voltage-dependent calcium channels undergo inhibitory modulation by G-protein activation, generally involving both kinetic slowing and steady-state inhibition. We have shown previously that the beta-subunit of neuronal calcium channels plays an important role in this process, because when it is absent, greater receptor-mediated inhibition is observed (). We therefore hypothesized that the calcium channel beta-subunits normally may occlude G-protein-mediated inhibition. Calcium channel beta-subunits bind to the cytoplasmic loop between transmembrane domains I and II of the alpha1-subunits (). We have examined the hypothesis that this loop is involved in G-protein-mediated inhibition by making chimeras containing the I-II loop of alpha1B or alpha1A inserted into alpha1E (alpha1EBE and alpha1EAE, respectively). This strategy was adopted because alpha1B (the molecular counterpart of N-type channels) and, to a lesser extent, alpha1A (P/Q-type) are G-protein-modulated, whereas this has not been observed to any great extent for alpha1E. Although alpha1B, coexpressed with alpha2-delta and beta1b transiently expressed in COS-7 cells, showed both kinetic slowing and steady-state inhibition when recorded with GTPgammaS in the patch pipette, both of which were reversed with a depolarizing prepulse, the chimera alpha1EBE (and, to a smaller extent, alpha1EAE) showed only kinetic slowing in the presence of GTPgammaS, and this also was reversed by a depolarizing prepulse. These results indicate that the I-II loop may be the molecular substrate of kinetic slowing but that the steady-state inhibition shown by alpha1B may involve a separate site on this calcium channel.
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K M
KM
Department of Pharmacology, Royal Free Hospital School of Medicine, London NW3 2PF, United Kingdom.
Stephens
G J
GJ
Berrow
N S
NS
Dolphin
A C
AC
D000818:Animals; D019556:COS Cells; D015220:Calcium Channels / Q000187:drug effects / Q000235:genetics* / Q000502:physiology*; D002678:Chimera; D004558:Electric Stimulation; D004594:Electrophysiology; D019204:GTP-Binding Proteins / Q000502:physiology*; D016244:Guanosine 5'-O-(3-Thiotriphosphate) / Q000494:pharmacology; D006706:Homeostasis; D007700:Kinetics; D011817:Rabbits; D051381:Rats
D015220:Calcium Channels; D016244:Guanosine 5'-O-(3-Thiotriphosphate); D019204:GTP-Binding Proteins