Inhibition of T cell proliferation by selective block of Ca(2+)-activated K(+) channels. (9/335)

T lymphocytes express a plethora of distinct ion channels that participate in the control of calcium homeostasis and signal transduction. Potassium channels play a critical role in the modulation of T cell calcium signaling, and the significance of the voltage-dependent K channel, Kv1.3, is well established. The recent cloning of the Ca(2+)-activated, intermediate-conductance K(+) channel (IK channel) has enabled a detailed investigation of the role of this highly Ca(2+)-sensitive K(+) channel in the calcium signaling and subsequent regulation of T cell proliferation. The role IK channels play in T cell activation and proliferation has been investigated by using various blockers of IK channels. The Ca(2+)-activated K(+) current in human T cells is shown by the whole-cell voltage-clamp technique to be highly sensitive to clotrimazole, charybdotoxin, and nitrendipine, but not to ketoconazole. Clotrimazole, nitrendipine, and charybdotoxin block T cell activation induced by signals that elicit a rise in intracellular Ca(2+)-e.g., phytohemagglutinin, Con A, and antigens such as Candida albicans and tetanus toxin in a dose-dependent manner. The release of IFN-gamma from activated T cells is also inhibited after block of IK channels by clotrimazole. Clotrimazole and cyclosporin A act synergistically to inhibit T cell proliferation, which confirms that block of IK channels affects the process downstream from T cell receptor activation. We suggest that IK channels constitute another target for immune suppression.  (+info)

Dihydropyridine and beta adrenergic receptor binding in dogs with tachycardia-induced atrial fibrillation. (10/335)

BACKGROUND: We have shown that rapid atrial activation, as occurs during atrial fibrillation (AF), reduces L-type Ca2+ current (ICa) and that this is the principal mechanism of the action potential duration and refractoriness changes that characterize tachycardia-induced atrial remodeling. The present study was designed to determine whether atrial tachycardia alters biochemical indices of the number of L-type Ca2+ channels and/or of the number and binding affinity of beta-adrenergic receptors. METHODS: In canine atrial sarcolemmal preparations, the number and binding affinity of dihydropyridine receptors were determined with the use of 3H-nitrendipine and that of beta-adrenergic receptors with 125I-iodocyanopindolol. Results were obtained with preparations from dogs paced at 400/min for 1 (P1, n = 20), 7 (P7, n = 9), and 42 (P42, n = 9) days, and compared with observations in sham-operated controls (P0, n = 14). RESULTS: Pacing reduced the Bmax of dihydropyridine receptors, from 157 +/- 18 fmol/mg (P0) to 116 +/- 9 fmol/mg (P1, P < 0.05), 100 +/- 14 fmol/mg (P7, P < 0.05) and 94 +/- 9 fmol/mg (P42, P < 0.01). The affinity of dihydropyridine receptors was unchanged, with the Kd averaging 711 +/- 102 pM. 656 +/- 74 pM, 633 +/- 155 pM and 585 +/- 92 pM in P0, P1, P7 and P42 dogs. Neither Bmax nor Kd of beta-adrenergic receptors was altered by rapid pacing. Values of Bmax of dihydropyridine receptors correlated with atrial ICa current density (r2 = 0.95) and ERP (r2 = 0.99). CONCLUSIONS: Rapid atrial activation results in downregulation in the number of dihydropyridine receptors without altering the number or affinity of beta-adrenergic receptors. The reductions in ICa that play an important role in the atrial electrical remodeling by which 'AF begets AF' appear to be due at least in part to a decrease in the number of L-type Ca2+ channels in cardiac cell membranes.  (+info)

Calcium channel activation and self-biting in mice. (11/335)

The L type calcium channel agonist (+/-)Bay K 8644 has been reported to cause characteristic motor abnormalities in adult mice. The current study shows that administration of this drug can also cause the unusual phenomenon of self-injurious biting, particularly when given to young mice. Self-biting is provoked by injecting small quantities of (+/-)Bay K 8644 directly into the lateral ventricle of the brain, suggesting a central effect of the drug. Similar behaviors can be provoked by administration of another L type calcium channel agonist, FPL 64176. The self-biting provoked by (+/-)Bay K 8644 can be inhibited by pretreating the mice with dihydropyridine L type calcium channel antagonists such as nifedipine, nimodipine, or nitrendipine. However, self-biting is not inhibited by nondihydropyridine antagonists including diltiazem, flunarizine, or verapamil. The known actions of (+/-)Bay K 8644 as an L type calcium channel agonist, the reproduction of similar behavior with another L type calcium channel agonist, and the protection afforded by certain L type calcium channel antagonists implicate calcium channels in the mediation of the self-biting behavior. This phenomenon provides a model for studying the neurobiology of this unusual behavior.  (+info)

Difference in mechanism between glyceraldehyde- and glucose-induced insulin secretion from isolated rat pancreatic islets. (12/335)

The effects of D-glyceraldehyde and glucose on islet function were compared in order to investigate the difference between them in the mechanism by which they induce insulin secretion. The stimulation of insulin secretion from isolated rat islets by 10 mM glyceraldehyde was not completely inhibited by either 150 microM diazoxide (an opener of ATP-sensitive K(+) channels) or 5 microM nitrendipine (an L-type Ca(2+)-channel blocker), whereas the stimulation of insulin secretion by 20 mM glucose was completely inhibited by either drug. The insulin secretion induced by glyceraldehyde was less augmented by 100 microM carbachol (a cholinergic agonist) than that induced by glucose. The stimulation of myo-inositol phosphate production by 100 microM carbachol was more marked in islets incubated with the hexose than with the triose. The content of glyceraldehyde 3-phosphate, a glycolytic intermediate, in islets incubated with glyceraldehyde was far higher than that in islets incubated with glucose, whereas the ATP content in islets incubated with the triose was significantly lower than that in islets incubated with the hexose. These results suggest that glyceraldehyde not only mimics the effect of glucose on insulin secretion but also has the ability to cause the secretion of insulin without the influx of Ca(2+ )through voltage-dependent Ca(2+) channels. The reason for the lower potency of the triose than the hexose in stimulating insulin secretion is also discussed.  (+info)

Tyrosine-kinase dependent TGF-beta and extracellular matrix expression by mechanical stretch in vascular smooth muscle cells. (13/335)

Vascular hypertrophy, which is characterized by proliferation of vascular smooth muscle cells (VSMC) and accumulation of extracellular matrix (ECM), is a major pathological change in blood vessels after chronic exposure to hypertension. Blood pressure is transmitted to the arterial walls and counterbalanced by mechanical stress, leading to stretching of circumferentially oriented VSMC, which may play some role in the pathogenesis of vascular hypertrophy. The present study was designed, therefore, to investigate the effect of mechanical stretch on the expression of ECM components and transforming growth factor-beta (TGF-beta), a potent stimulator for ECM production, and to examine the signal transduction mechanisms of the induction of TGF-beta in cultured rat VSMC. VSMC were subjected to cyclic stretch to provide a maximal elongation of 20% at a rate of 60 cycles per minute for up to 24 h. Mechanical stretch stimulated TGF-beta1 mRNA expression in a time- and elongation-dependent manner. Indeed, the secretion of TGF-beta proteins into the culture media was increased after stretch. Stretch also stimulated mRNA expression of the ECM components, type I and type IV collagen, and fibronectin, which was largely inhibited by addition of neutralizing antibody against TGF-beta. The tyrosine kinase inhibitors genistein and herbimycin A blocked the induction of TGF-beta1 and type I collagen by stretch, while protein kinase C inhibitors, the calcium channel blockers nitrendipine and gadolinium, or Ca removal from the media had no effect. These results suggest that stretch-induced, tyrosine kinase-mediated autocrine/paracrine production of TGF-8 may play a critical role in the progression of vascular remodeling associated with high blood pressure.  (+info)

Functional alteration of dihydropyridine-sensitive Ca(2+) channels in the adrenal glomerulosa of pregnant rats. (14/335)

Our previous work on aldosterone secretion suggested that dihydropyridine-sensitive calcium channels, one type of voltage-dependent calcium channels (VDCC), are functionally impaired in adrenal capsule preparations from the pregnant rat. The aim of this study was to determine whether, during pregnancy, the density and/or activity of these channels is altered in the adrenal zona glomerulosa. These VDCC measured with [(3)H]nitrendipine binding were not different between membrane preparations of nonpregnant and pregnant rats. Western blots were performed using two different antibodies, a polyclonal (PcAb) directed against the alpha(1)-subunit of VDCC and a monoclonal (McAb) that recognizes an intracellular domain of that protein. McAb immunoreactivity showed a significant decrease in preparations from pregnant rats, whereas no difference was observed with PcAb. VDCC activity was estimated by (45)Ca(2+) uptake in isolated adrenal cortex and by intracellular calcium concentration ([Ca(2+)](i)) in adrenal glomerulosa cells with the Ca(2+) probe fura PE3. These measurements revealed that KCl stimulation produced greater Ca(2+) influx in nonpregnant than in pregnant rats. Nifedipine (a blocker of VDCC) inhibited this stimulation only in nonpregnant rats, whereas BAY K 8644 (an activator of VDCC) increased Ca(2+) influx in pregnant rats only. These data suggest that, during pregnancy, the altered regulation of calcium homeostasis in adrenal glomerulosa is linked to a conformational alteration of VDCC.  (+info)

CaV2.2 and CaV2.3 (N- and R-type) Ca2+ channels in depolarization-evoked entry of Ca2+ into mouse sperm. (15/335)

As sperm prepare for fertilization, surface Ca(2+) channels must open to initiate required, Ca(2+)-mediated events. However, the molecular identity and functional properties of sperm Ca(2+) channels remain uncertain. Here, we use rapid local perfusion and single-cell photometry to examine the kinetics of calcium responses of mouse sperm to depolarizing stimuli. The linear rise of intracellular [Ca(2+)] evoked by approximately 10-s applications of an alkaline high [K(+)] medium directly reports activity of voltage-gated Ca(2+) channels. Little response occurs if external Ca(2+) is removed or if external or internal pH is elevated without depolarization. Responses are inhibited 30-40% by 30-100 micrometer Ni(2+) and more completely by 100-300 micrometer Cd(2+). They resist the dihydropyridines nitrendipine and PN200-110, but 1-10 micrometer mibefradil inhibits reversibly. They also resist the venom toxins calciseptine, omega-conotoxin MVIIC, and kurtoxin, but omega-conotoxin GVIA (5 micrometer) inhibits approximately 50%. GVIA also partially blocks transient, low voltage activated Ca(2+) currents of patch-clamped spermatids. Differential sensitivity of sperm responses to Ni(2+) and Cd(2+) and partial blockade by GVIA indicate that depolarization opens at least two types of voltage-gated Ca(2+) channels in epididymal sperm examined prior to capacitation. Involvement of a previously undetected Ca(V)2.2 (N-type) channel, suggested by the action of GVIA, is substantiated by immunodetection of Ca(2+) channel alpha(1B) subunits in sperm and sperm extracts. Resistance to dihydropyridines, calciseptine, MVIIC, and kurtoxin indicates that Ca(V)1, Ca(V)2.1, and Ca(V)3 (L-, P/Q-, and T-type) channels contribute little to this evoked response. Partial sensitivity to 1 micrometer mibefradil and an enhanced sensitivity of the GVIA-resistant component of response to Ni(2+) suggest participation of a Ca(V)2.3 (R-type) channel specified by previously found alpha(1E) subunits. Our examination of depolarization-evoked Ca(2+) entry indicates that mature sperm possess a larger palette of voltage-gated Ca(2+) channels than previously thought. Such diversity may permit specific responses to multiple cues encountered on the path to fertilization.  (+info)

Dihydropyridine-induced Ca2+ release from ryanodine-sensitive Ca2+ pools in human skeletal muscle cells. (16/335)

Dihydropyridines (DHPs) are widely used antihypertensive drugs and inhibit excitation-contraction (E-C) coupling in vascular smooth muscle and in myocardial cells by antagonizing L-type Ca2+ channels (DHP receptors). However, contradictory reports exist about the interaction of the DHP with the skeletal muscle isoform of the DHP receptor and E-C coupling in skeletal muscle cells. Using the intracellular fluorescent Ca2+ indicator fura-2, an increase in [Ca2+]i was observed after extracellular application of nifedipine to cultured human skeletal muscle cells. The rise in [Ca2+]i was dose dependent with a calculated EC50 of 614 +/- 96 nM nifedipine and a maximum increment in [Ca2+]i of 80 +/- 3.2 nM. Similar values were obtained with nitrendipine. This effect of DHPs was restricted to differentiated skeletal muscle cells and was not seen in non-differentiated cells or in PC12 cells. In spite of the observed increase in [Ca2+]i, whole-cell patch clamp experiments revealed that 10 microM nifedipine abolished inward Ba2+ currents through L-type Ca2+ channels completely. Similar to nifedipine, (+/-)Bay K 8644, an agonist of the L-type Ca2+ channel, also increased [Ca2+]i. This effect could not be enhanced by further addition of nifedipine, suggesting that both DHPs act via a common signalling pathway. Based on the specific mechanism of the skeletal muscle E-C coupling, we propose the stabilization of a conformational state of the DHP receptor by DHPs, which is sufficient to activate the ryanodine receptor.  (+info)