Molecular biology of the calcium antagonist receptor.

The calcium channel plays a key role in controlling many physiological processes in the body. Drugs that block the calcium channel have proven clinically effective for the treatment of a multitude of cardiovascular disorders. The elucidation of the precise mechanism of action of these drugs involves cloning the calcium channels on which they act. Genetic manipulation of these cloned channels is beginning to reveal the binding sites for the calcium channel blocking drugs and may lead to the development of more specific agents.

Characterization of beta subunit modulation of a rabbit cardiac L-type Ca2+ channel alpha 1 subunit as expressed in mouse L cells.

Functional properties of a rabbit cardiac alpha 1 Ca2+ channel subunit (CARD alpha 1) were investigated using the patch-clamp technique in mouse L cells, a recipient cell line which is devoid of any Ca2+ channel subunits. Cell lines resulting from stable transfection of the CARD alpha 1 subunit as well as in coexpression with a beta subunit (CARD alpha 1 beta) derived from skeletal muscle (SKM beta) were characterized. The results show that while the CARD alpha 1-Ca2+ channel activity is negligible, the Ba2+ current density is dramatically increased in the presence of beta subunit (approximately 20-fold). CARD alpha 1- and CARD alpha 1 beta-Ba2+ currents were both sensitive to the 1,4-dihydropyridine (DHP) agonist, Bay K 8644 (5- to 8-fold increase). Activation kinetics of CARD alpha 1- and CARD alpha 1 beta-Ba2+ currents were comparable. The inactivation time-course was faster (3- to 4-fold) for CARD alpha 1 beta-Ba2+ currents. We conclude that the main role of the beta subunit in heart is to modulate the L-type current density and present several lines of evidence that SKM alpha 1 and CARD alpha 1 are differentially regulated by the beta subunit.

Evidence for the existence of a cardiac specific isoform of the alpha 1 subunit of the voltage dependent calcium channel.

Biochemical, pharmacological and electrophysiological evidence implies the existence of tissue specific isoforms of the L-type VDCC. The alpha 1 and alpha 2 subunits of the skeletal muscle calcium channel have been previously cloned and their amino acid sequence deduced. Here we report the isolation and sequencing of a partial cDNA that encodes a heart specific isoform of the alpha 1 subunit. The amino acid sequence deduced from this part cDNA clone shows 64.7% similarity with the skeletal muscle alpha 1 subunit. Northern analysis reveals 2 hybridizing bands, 8.5 and 13 kb, in contrast to one 6.5 kb band in the skeletal muscle. Selective inhibition of mRNA expression in Xenopus oocytes by complementary oligodeoxy-nucleotides derived from the heart clone provides further evidence that the cDNA corresponds to an essential component of the VDCC. These data further support the existence of tissue-specific isoforms of the L-type VDCC.

Native-type DHP-sensitive calcium channel currents are produced by cloned rat aortic smooth muscle and cardiac alpha 1 subunits expressed in Xenopus laevis oocytes and are regulated by alpha 2- and beta-subunits.

Native tissue-like L-type voltage-dependent calcium channels (L-VDCC's) were expressed by in vitro transcribed cRNA injection of rat aorta or rabbit cardiac alpha 1 subunit into Xenopus laevis oocytes. Co-injection of VSM-alpha 1 with the cloned skeletal muscle beta-subunit (SK-beta) of the L-type VDCC significantly increased the expressed peak current amplitude without significant changes in kinetics. Similar results were obtained by co-injection of cardiac alpha 1 (DSHT-alpha 1) the cloned skeletal alpha 2-subunit (SK-alpha 2) or with SK-beta. The oocytes co-expressing cRNA's retained L-type VDCC pharmacology.

Toward an understanding of the dihydropyridine-sensitive calcium channel.

The dihydropyridine-sensitive calcium channel continues to fascinate scientific investigators. Each new discovery leads to more complexity. Further work elucidating the molecular structures and functions of the various isoforms of the channel will lead us to a better understanding of its nature. We are well on our way towards understanding the molecular structure and mechanisms involved in calcium permeability, and the coming decade promises to reveal numerous breakthroughs in our understanding of this channel.

Ethnobotany in the search for vasoactive herbal medicines.

Plant samples derived from ethno-directed and random collections were screened to determine their effect on pre-contracted rat aortic tissue. Of the 31 ethno-directed species, four were found to be potent relaxants of vascular smooth muscle. The vasoactive species were Chamguava gentlei, Alseis yucatanensis, Licaria peckii and Nectandra salicifolia. None of the 32 randomly collected samples produced a relaxation response. These data support the hypothesis that ethno-directed collection is a more efficient means of drug discovery than random plant screens.