8-bromocyclic-gmp and 1-4-dihydropyridine

Smoking is a major health hazard with proven deleterious effects on the cerebral circulation, including a decrease in cerebral blood flow and a high risk for stroke. To elucidate cellular mechanisms for the vasoconstrictive and pathological effects of nicotine, we used a nystatin-perforated patch-clamp technique to study Ca(2+) channels and Ca(2+)-activated K(+) (BK) channels in smooth muscle cells isolated from cerebral lenticulostriate arterioles of rats chronically exposed to nicotine (4.5 mg/kg per day of nicotine free base, 15 to 22 days via osmotic minipump). Two major effects were observed in cells from nicotine-treated animals compared with controls. First, Ca(2+) channels were upregulated (0.48+/-0.03 pS/pF [20 cells] versus 0.35+/-0.01 pS/pF [31 cells], P:<0.005) and BK channels were downregulated (12+/-3 pA/pF [14 cells] versus 34+/-7 pA/pF [14 cells], P:<0.05), mimicking the effect of an apparent decrease in bioavailability of endogenous NO. Second, normal downregulation of Ca(2+) channels by exogenous NO (sodium nitroprusside [SNP], 100 nmol/L) and cGMP (8-bromo-cGMP, 0.1 mmol/L) was absent, whereas normal upregulation of BK channels by these agents was preserved, suggesting block of NO signaling downstream of cGMP-dependent protein kinase. In pial window preparations, chronic nicotine blunted NO-induced vasodilation of pial vessels and the increase in cortical blood flow measured by laser-Doppler flowmetry, demonstrating the importance of Ca(2+) channel downregulation in NO-induced vasorelaxation. These findings elucidate a new pathophysiological mechanism involving altered Ca(2+) homeostasis in cerebral arterioles that may predispose to stroke.

Topics: 3-Pyridinecarboxylic acid, 1,4-dihydro-2,6-dimethyl-5-nitro-4-(2-(trifluoromethyl)phenyl)-, Methyl ester; Animals; Arterioles; Barium; Calcium; Calcium Channels; Calcium Channels, L-Type; Cerebral Arteries; Cerebrovascular Circulation; Cyclic GMP; Dihydropyridines; Electric Stimulation; Female; Membrane Potentials; Muscle, Smooth, Vascular; Nicotine; Nifedipine; Nitric Oxide; Nitroprusside; Potassium Channels; Rats; Rats, Inbred WKY; Signal Transduction; Time Factors; Tyrphostins

Pretreatment of chromaffin cells with the permeable analogue of cGMP, 8-Br-cGMP (100 microM), leads to a reduction (35%) of depolarization-evoked intracellular calcium concentration ([Ca2+]i) increases. There is evidence that bovine adrenal chromaffin cells are provided with both dihydropyridine-sensitive and -resistant voltage-sensitive Ca2+ influx pathways. Combined incubations with nifedipine 10 microM and 8-Br-cGMP reduced KCl-evoked intracellular Ca2+ concentration to a greater extent that each compound separately. Moreover, 8-Br-cGMP failed to affect the [Ca2+]i transient induced by the L-type Ca2+ channel agonist Bay K 8644 (1 microM) under conditions of low depolarization. Neomycin (0.2 mM) and omega-AgaToxin-IVA (AgTx) (1 microM) inhibited the calcium transient to a similar extent, and this inhibition was not enhanced by the presence of 8-Br-cGMP. It is concluded that 8-Br-cGMP modulated the dihydropyridine-insensitive Ca2+ influx pathway in the chromaffin cell.

Topics: 3-Pyridinecarboxylic acid, 1,4-dihydro-2,6-dimethyl-5-nitro-4-(2-(trifluoromethyl)phenyl)-, Methyl ester; Adrenal Medulla; Animals; Bucladesine; Calcium; Calcium Channels; Cattle; Cells, Cultured; Cyclic GMP; Dihydropyridines; Neomycin; Nerve Tissue Proteins; omega-Agatoxin IVA; omega-Conotoxin GVIA; Peptides; Potassium Chloride; Second Messenger Systems; Spider Venoms