Fiber-FISH analysis of the 3'-terminal region of the human L-type Ca2+ channel alpha 1C subunit gene

Human L-type Ca2+ channel alpha 1C subunit gene (CACNL1A1) maps to the distal region of chromosome 12p13, and is composed of approximately 50 exons spanning over 150 kb of the human genome as estimated by restriction map analysis. However, the structure and the total length of the 3'-end of the gene is not clear because the size of several big introns remains unknown. Here the fiber-FISH technique was used to determine the relative order and size of eight partial genomic DNA clones from the central and 3'-terminal regions of CACNL1A1. The total physical distance of this region, including the size and gap distances between the clones were re-estimated. The results show that the physical order of the tested clones was 5'-g14-5 > g12-2 > g10-8 > g4-5 > g16-7 > g8-3 > g12-5 > g6-20-3'. Their individual sizes vary between 6.7 and 21.9 kb. Clones g6-20 and g12-5, both containing repetitive exon 45/46-like element, were found to be located within 59.1 kb downstream of g8-3 containing earlier identified polyadenylation site, i.e. 229.5 kb away from clone g14-5 (exons 10, 11). The possible implications of this structural complexity is discussed.     

Ca(2+) sensors of L-type Ca(2+) channel

Ca(2+)-induced inactivation of L-type Ca(2+) is differentially mediated by two C-terminal motifs of the alpha(1C) subunit, L (1572-1587) and K (1599-1651) implicated for calmodulin binding. We found that motif L is composed of a highly selective Ca(2+) sensor and an adjacent Ca(2+)-independent tethering site for calmodulin. The Ca(2+) sensor contributes to higher Ca(2+) sensitivity of the motif L complex with calmodulin. Since only combined mutation of both sites removes Ca(2+)-dependent current decay, the two-site modulation by Ca(2+) and calmodulin may underlie Ca(2+)-induced inactivation of the channel.     

Histochemical and immunohistochemical identification of endocrine cells in the internal ear

The authors describe the results of the study of rabbit decalcified temporal bones and guinea-pig membranous cochlea treated according to the Grimelius argyrophil method and immunohistochemical technique with the use of a number of specific antisera. The study revealed the cells of the diffuse endocrine system (APUD system) in various parts of the internal ear: in the main membrane, receptor elements of the spiral chorda, and spiral ligament. Serotonin and melatonin were identified in the cells of the APUD system. The data obtained may form a theoretical basis of a basically new trend in the study of the function of the internal ear in health and disease.     

Photoactivated radioactive analogs of calcium antagonists of 1,4-dihydropyridine series: synthesis and covalent labeling of the receptor in rabbit skeletal muscle

Two novel radioactive 1.4-dihydropyridine derivatives were synthesized and their properties were shown to be characteristic for calcium antagonists of nifedipine series. Photoactivable o- and p-azidobenzoyl groups as well as one or two tritium atoms were introduced into the reactive site of these compounds. The binding of molecular probes obtained with rabbit skeletal muscle membranes was characterized by a high level of specificity, the affinity constant being about 10(-9) M. Upon UV-light photoactivation, the radioactive probes selectively incorporated into skeletal muscle membrane protein (Mr approximately 136 kD).