High-fat diet elevates blood pressure and cerebrovascular muscle Ca(2+) current.

Dietary fat contributes to the elevation of blood pressure and increases the risk of stroke and coronary artery disease. Previous observations have shown that voltage-gated Ca(2+) current density is significantly increased in hypertension and can be affected by free fatty acids (FAs). We hypothesized that a diet of elevated fat level would lead to an increase in blood pressure, an elevation of L-type Ca(2+) current, and an increase in saturated FA content in vascular smooth muscle cell membranes. Male Osborne-Mendel rats were fed normal rat chow or a high-fat diet (Ob/HT group) for 8 weeks. Blood pressures in the Ob/HT group increased moderately from 122.5+/-0.7 to 134.4+/-0.8 mm Hg (P<0.05, n=26). Voltage-clamp examination of cerebral arterial cells revealed significantly elevated L-type Ca(2+) current density in the Ob/HT group. Voltage-dependent inactivation of the Ob/HT L-type channels was significantly delayed. Total serum FA contents were significantly elevated in the Ob/HT group, and HPLC analyses of fractional pools of FAs from segments of abdominal aorta revealed that arachidonic acid levels were elevated in the phospholipid fraction in Ob/HT. No differences in vascular membrane cholesterol contents were noted. Plasma cholesterol was significantly elevated in portal venous and cardiac blood samples from Ob/HT rats. These findings suggest that an elevation of plasma FAs may contribute to the development of hypertension via a process involving the elevation of Ca(2+) current density and an alteration of channel kinetics in the vascular smooth muscle membrane.

Ceramide-induced vasorelaxation: An inhibitory action on protein kinase C.

Experiments were designed to examine the role of sphingosine, PP2A phosphatases, and protein kinase C (PKC) inhibition in mediating the vasodilatory effects of ceramide in rat thoracic aorta. Sphingosine did not cause vasorelaxation, and oleoylethanol-amine, a ceramidase inhibitor, did not affect sphingomyelinase-induced relaxation. Okadaic acid potentiated the relaxation response to ceramide. These observations rule out involvement of sphingosine and PP2A phosphatases in mediating ceramide-induced relaxation. Sphingomyelinase attenuated contractile and single-cell intracellular calcium responses to phorbol ester. Chelerythrine incubation potentiated the relaxation response to ceramide. These observations support a role for PKC inhibition in mediating the vasodilatory effects of ceramide.

Linoleic acid induces relaxation and hyperpolarization of the pig coronary artery.

Linoleic acid, a polyunsaturated C18 fatty acid, is one of the major fatty acids in the coronary arterial wall. Although diets rich in linoleic acid reduce blood pressure and prevent coronary artery disease in both humans and animals, very little is known about its mechanism of action. We believed that its beneficial effects might be mediated by changes in vascular tone. We investigated whether linoleic acid induces relaxation of porcine coronary artery rings and the mechanism involved in this process. Linoleic acid and two of its metabolites, 13-hydroxyoctadecadienoic acid (13-HODE) and 13-hydroperoxyoctadecadienoic acid (13-HPODE), induced dose-dependent relaxation of prostaglandin (PG) F2alpha-precontracted rings that was not affected by indomethacin (10[-5] mol/L), a cyclooxygenase inhibitor, or cinnamyl-3,4-dihydroxy-alpha-cyanocinnamate (CDC; 10[-5] mol/L), a lipoxygenase inhibitor. Removal of endothelial cells had no effect on vasorelaxation, suggesting a direct effect on the vascular smooth muscle cells (VSMC). When rings were contracted with KCl, linoleic acid failed to induce relaxation. Although tetrabutylammonium (5 x 10[-3] mol/L), a nonselective K+ channel blocker, slightly inhibited the relaxation caused by linoleic acid, glibenclamide (10[-6] mol/L), an ATP-sensitive K+ channel blocker, and charybdotoxin (7.5x10[-8] mol/L) or tetraethylammonium (5x10[-3] mol/L), two different Ca2+-activated K+ channel blockers, had no effect. However, relaxation was completely blocked by ouabain (5x10[-7] mol/L), a Na+/K+-ATPase inhibitor, or by a K+-free solution. In addition, linoleic acid (10[-6] mol/L) caused sustained hyperpolarization of porcine coronary VSMC (from -49.5+/-2.0 to -60.7+/-4.2 mV), which was also abolished by ouabain. We concluded that linoleic acid induces relaxation and hyperpolarization of porcine coronary VSMC via a mechanism that involves activation of the Na+/K+-ATPase pump.

Calcium handling by vascular myocytes in hypertension.

Calcium ions (Ca2+) trigger the contraction of vascular myocytes and the level of free intracellular Ca2+ within the myocyte is precisely regulated by sequestration and extrusion mechanisms. Extensive evidence indicates that a defect in the regulation of intracellular Ca2+ plays a role in the augmented vascular reactivity characteristic of clinical and experimental hypertension. For example, arteries from spontaneously hypertensive rats (SHR) have an increased contractile sensitivity to extracellular Ca2+ and intracellular Ca2+ levels are elevated in aortic smooth muscle cells of SHR. We hypothesize that these changes are due to an increase in membrane Ca2+ channel density and possibly function in vascular myocytes from hypertensive animals. Several observations using various experimental approaches support this hypothesis: 1) the contractile activity in response to depolarizing stimuli is increased in arteries from hypertensive animals demonstrating increased voltage-dependent Ca2+ channel activity in hypertension; 2) Ca2+ channel agonists such as Bay K 8644 produce contractions in isolated arterial segments from hypertensive rats and minimal contraction in those from normotensive rats; 3) intracellular Ca2+ concentration is abnormally increased in vascular myocytes from hypertensive animals following treatment with Ca2+ channel agonists and depolarizing interventions, and 4) using the voltage-clamp technique, the inward Ca2+ current in arterial myocytes from hypertensive rats is nearly twice as large as that from myocytes of normotensive rats. We suggest that an alteration in Ca2+ channel function and/or an increase in Ca2+ channel density, resulting from increased channel synthesis or reduced turnover, underlies the increased vascular reactivity characteristic of hypertension.

Calcium handling by vascular myocytes in hypertension

Calcium ions (Ca2+) trigger the contraction of vascular myocytes and the level of free intracellular Ca2+ within the myocyte is precisely regulated by sequestration and extrusion mechanisms. Extensive evidence indicates that a defect in the regulation of intracellular Ca2+ plays a role in the augmented vascular reactivity characteristic of clinical and experimental hypertension. For example, arteries from spontaneously hypertensive rats (SHR) have an increased contractile sensitivity to extracellular Ca2+ and intracellular Ca2+ levels are elevated in aortic smooth muscle cells of SHR. We hypothesize that these changes are due to an increase in membrane Ca2+ channel density and possibly function in vascular myocytes from hypertensive animals. Several observations using various experimental approaches support this hypothesis: 1) the contractile activity in response to depolarizing stimuli is increased in arteries from hypertensive animals demonstrating increased voltage-dependent Ca2+ channel activity in hypertension; 2) Ca2+ channel agonists such as Bay K 8644 produce contractions in isolated arterial segments from hypertensive rats and minimal contraction in those from normotensive rats; 3) intracellular Ca2+ concentration is abnormally increased in vascular myocytes from hypertensive animals following treatment with Ca2+ channel agonists and depolarizing interventions, and 4) using the voltage-clamp technique, the inward Ca2+ current in arterial myocytes from hypertensive rats is nearly twice as large as that from myocytes of normotensive rats. We suggest that an alteration in Ca2+ channel function and/or an increase in Ca2+ channel density, resulting from increased channel synthesis or reduced turnover, underlies the increased vascular reactivity characteristic of hypertension.

The effects of cyclopiazonic acid on intracellular Ca2+ in aortic smooth muscle cells from DOCA-hypertensive rats.

We tested the hypothesis that cyclopiazonic acid (CPA), an inhibitor of the sarcoplasmic reticulum (SR) Ca(2+)-ATPase, increases intracellular Ca2+ concentration ([Ca2+]) in aortic myocytes and that the increase in [Ca2+]i is higher in aortic cells from deoxycorticosterone acetate (DOCA)-hypertensive rats. Male Sprague-Dawley rats, 250-300 g, underwent uninephrectomy, received a silastic implant containing DOCA (200 mg/kg) and had free access to water supplemented with 1.0% NaCl and 0.2% KCl. Control rats were also uninephrectomized, received normal tap water, but no implant. Intracellular Ca2+ measurements were performed in aortic myocytes isolated from normotensive (Systolic blood pressure = 120 +/- 3 mmHg; body weight = 478 +/- 7 g, N = 7) and DOCA-hypertensive rats (195 +/- 10 mmHg; 358 +/- 16 g, N = 7). The effects of CPA on resting [Ca2+]i and on caffeine-induced increase in [Ca2+]i after [Ca2+]i depletion and reloading were compared in aortic cells from DOCA and normotensive rats. The phasic increase in [Ca2+]i induced by 20 mM caffeine in Ca(2+)-free buffer was significantly higher in DOCA aortic cells (329 +/- 36 nM, N = 5) compared to that in normotensive cells (249 +/- 16 nM, N = 7, P < 0.05). CPA (3 microM) inhibited caffeine-induced increases in [Ca2+]i in both groups. When the cells were placed in normal buffer (1.6 mM Ca2+, loading period), after treatment with Ca(2+)-free buffer (depletion period), an increase in [Ca2+]i was observed in DOCA aortic cells (45 +/- 11 nM, N = 5) while no changes were observed in normotensive cells. CPA (3 microM) potentiated the increase in [Ca2+]i (122 +/- 30 nM, N = 5) observed in DOCA cells during the loading period while only a modest increase in [Ca2+]i (23 +/- 10 nM, N = 5) was observed in normotensive cells. CPA-induced increase in [Ca2+]i did not occur in the absence of extracellular Ca2+ or in the presence of nifedipine. These data show that CPA induces Ca2+ influx in aorta from both normotensive and DOCA-hypertensive rats. However, the increase in [Ca2+]i is higher in DOCA aortic cells possibly due to an impairment in the mechanisms that control [Ca2+]i. The large increase in [Ca2+]i in response to caffeine in DOCA cells probably reflects a greater storage of Ca2+ in the SR.

The effects of cyclopiazonic acid on intracellular Ca2+ in aortic smooth muscle cells from DOCA-hypertensive rats

We tested the hypothesis that cyclopiazonic acid (CPA), an inhibitor of the sarcoplasmic reticulum (SR) Ca2+ -ATPase, increases intracellular Ca2+ concentration ([Ca2+]i) in aortic myocytes and that the increase in [Ca2+]i is higher in aortic cells from deoxycorticosterone acetate (DOCA)-hypertensive rats. Male Sprague-Dawley rats, 250-300 g, underwent uninephrectomy, received a silastic implant containing DOCA (200 mg/kg) and had free access to water supplemented with 1.0 per cent NaCl and 0.2 per cent KCl. Control rats were also uninephrectomized, received normal tap water, but no implant. Intracellular Ca2+ measurements were performed in aortic myocytes isolated from normotensive (Systolic blood pressure = 120 + 3 mmHg; body weight = 478 ñ 7 g, N = 7) and DOCA-hypertensive rats (195 ñ 1O mmHg; 358 ñ 16 g, N = 7). The effects of CPA on resting [Ca2+]i and on caffeine-induced increase in [Ca2+]i after [Ca2+]i depletion and reloading were compared in aortic cells from DOCA and normotensive rats. The phasic increase in [Ca2+]i induced by 20 mM caffeine in Ca2+ -free buffer was significantly higher in DOCA aortic cells (329 ñ 36 nM, N = 5) compared to that in normotensive cells (249 ñ 16 nM, N = 7, P<0.05). CPA (3 muM) inhibited caffeine-induced increases in [Ca2+]i in both groups. When the cells were placed in normal buffer (1.6 mM Ca2+, loading period), after treatment with Ca2+ -free buffer (depletion period), an increase in [Ca2+]i was observed in DOCA aortic cells (45 ñ 11 nM, N = 5) while no changes were observed in normotensive cells. CPA (3 muM) potentiated the increase in [Ca2+]i (l22 ñ 3O nM, N = 5) observed in DOCA cells during the loading period while only a modest increase in [Ca2+]i, (23 ñ 10 nM, N = 5) was observed in normotensive cells. CPA-induced increase in [Ca2+]i did not occur in the absence of extracellular Ca2+ or in the presence of nifedipine. These data show that CPA induces Ca2+ influx in aorta from both normotensive and DOCA-hypertensive rats. However, the increase in [Ca2+]i is higher in DOCA aortic cells possibly due to an impairment in the mechanisms that control [Ca2+]i. The large increase in [Ca2+]i in response to caffeine in DOCA cells probably reflects a greater storage of Ca2+ in the SR.

Isoflurane reduces K+ current in single smooth muscle cells of guinea pig portal vein.

Volatile anesthetics vasodilate in part by direct action on vascular smooth muscle. Isoflurane-induced relaxation of portal vein smooth muscle involves alteration of membrane ionic currents that control cell excitability and contraction. Whole cell voltage clamp technique was used to examine outward Ca(2+)-activated K+ current (IK,Ca) in guinea pig portal vein cells. Isoflurane caused a concentration-dependent reduction in IK,Ca at steady-state conditions but had no significant effect on resting potential. Isoflurane transiently potentiated IK,Ca by a mechanism that may partly involve Ca2+ release from intracellular storage sites. The depression of IK,Ca by isoflurane may occur by direct action on the channel protein or on the lipid environment of the channel to alter conductance or kinetic properties. Since isoflurane reduces IK,Ca coincident with suppression of Ca2+ channel current, it was concluded that the depression of IK,Ca by isoflurane is of secondary importance to reduction in inward Ca2+ channel current. Potentiation of IK,Ca may preclude significant membrane activation during the onset of isoflurane's action.

Calcium channel activity increased by plasma from ischemic hindlimbs of rats: role of an endogenous NO synthase inhibitor.

We tested the hypothesis that an endogenous nitric oxide synthase (NOS) inhibitor released from ischemic hindlimbs increases the activity of calcium channels in vascular smooth muscle, thus contributing to the increased contractile response to calcium agonists. Hindlimb ischemia was generated in rats by infrarenal aortic cross clamping for 5 h, after which plasma was obtained from femoral vein blood. Incubating naive aortic rings (endothelium intact) for 2 h in plasma collected from ischemic rats significantly reduced relaxation to acetylcholine in precontracted rings and increased contraction to the calcium channel agonist, BAY K 8644. However, in isolated smooth muscle cells (without endothelium) loaded with fura-2, no difference was noted in BAY K 8644-stimulated intracellular calcium concentration. The contractile responses to sodium fluoride, serotonin, and calcium ionophore A23187 were not different in either ischemic or control plasma-incubated rings. The augmentation of the contractile response to BAY K 8644 was significantly inhibited by nitroglycerin (10-8 M) and by exposure to calcium-free solution. N omega-nitro-L-arginine (without plasma incubation)-pretreated rings also demonstrated hyperresponsiveness to BAY K 8644. The increase in responsiveness to BAY K 8644 exhibited a negative correlation with the maximal relaxation to acetylcholine (r = -0.99), suggesting that the apparent increase in activity of calcium channels is mediated through inhibition of nitric oxide by an endogenous NOS inhibitor on endothelium.

Interactions of ethanol and quinidine on contractility and myocyte action potential in the rat ventricle.

The combined effects of ethanol and quinidine on cardiac electromechanical coupling are unknown, but both drugs affect cardiac conduction and can cause myocardial depression. Isolated left ventricular papillary and ventricular myocytes were used to assess the combined effects of quinidine and ethanol on the electrophysiologic and mechanical properties of rat myocardium. The combination of quinidine (1-300 microM) and ethanol (120-240 mg/dL) depressed active papillary muscle tension within the clinically useful concentration range. In electrophysiologic studies of isolated ventricular myocytes, quinidine prolonged the action potential duration at 50% (APD50) and 90% (APD90) repolarization, the absolute refractory period, and the relative refractory period, but decreased the maximum rate of change of depolarization in phase 0 (Vmax). When cells were exposed to ethanol (240 mg/dL) and quinidine (1.5 microM) together, a significant decrease in the quinidine-induced prolongation of the absolute refractory and relative refractory periods was seen. Additional changes in action potential parameters from the quinidine values included slight reductions in Vmax and in APD50 and APD90, but these reductions were not consistently displayed, nor were they statistically significant.

Isoflurane reduces Ca++ channel current and accelerates current decay in guinea pig portal vein smooth muscle cells.

The volatile anesthetic isoflurane (ISO) can exert significant inhibitory effects on hemodynamics and organ perfusion. It was hypothesized that venodilation during ISO exposure was mediated through a reduction in inward Ca++ channel current. By using the whole cell mode of single cell patch clamp technique, the action of ISO on L-type Ca++ channel current was examined. ISO caused a concentration-dependent reduction in maximal Ca++ channel current and shifted the current activation to more negative potentials. ISO, 3%, shifted the voltage for half-maximal channel inactivation from -21.9 to -33.4 mV. Measurement of time constants for inward current activation and inactivation revealed that the inactivation phase exhibited a concentration-dependent reduction over the range of 0.36 to 3% ISO. The decrease in the inactivation time constant is consistent with accelerated inactivation of the L-channel current. Estimated binding constants for ISO to resting or inactivated channels suggest absence of state-dependent inhibition of channel conductance. The venodilatory effects of ISO are due in part to limitation of Ca++ influx which may secondarily affect Ca(++)-dependent mechanisms such as the outward Ca(++)-activated K+ channel population. Reduction in current by ISO may occur through receptor-independent mechanisms and may involve perturbation of the sarcolemmal lipid bilayer.

Calcium current in smooth muscle cells from normotensive and genetically hypertensive rats.

Genetic hypertension results from numerous phenotypic expressions. We hypothesized that increased calcium current in vascular smooth muscle of genetically hypertensive animals is partly responsible for observed increases in agonist sensitivity, contractility, and calcium influx. Using adult, spontaneously hypertensive stroke-prone rats (SHRSP) and normotensive Wistar-Kyoto (WKY) controls from an inbred colony, we characterized calcium current in smooth muscle cells isolated from cerebral arteries. Calcium current in WKY cells reached a maximum of -27.7 +/- 2.7 pA (n = 32) at +20 mV. Peak inward current at +20 mV in SHRSP cells had a mean amplitude of -44.4 +/- 3.0 pA (n = 72, P < .05). SHRSP cells exhibited a higher calcium current density. Maximal inward current normalized to cell capacitance yielded mean values of 2.07 +/- 0.11 pA/pF for WKY (n = 32) and 2.80 +/- 0.12 pA/pF (n = 79) for SHRSP (P < .05) cells. Transient-type Ca2+ channel current had the same magnitude and current-voltage relation in both cell types, giving an L-type/T-type ratio of 3.85 for WKY and 6.25 for SHRSP cells. The voltage-dependent inactivation curve for SHRSP calcium current was shifted to the right only over the range of -50 to -30 mV, but the half-maximal inactivation voltages and Boltzmann coefficients were not significantly different between cell types. Increased calcium inward current in this model of genetic hypertension could account in part for altered calcium homeostasis and increased vascular reactivity, contributing to hypertension and vasospasm.

Effects of isoflurane and enflurane on intracellular Ca2+ mobilization in isolated cardiac myocytes.

BACKGROUND: Enflurane and isoflurane may reduce cardiac contractility by altering mobilization and clearance of intracellular Ca2+ (Ca2+i). It was hypothesized that the negative inotropic actions of these agents involve limiting both membrane Ca2+ entry and altering intracellular Ca2+ release. METHODS: The Ca2+i transients in rat ventricular myocytes loaded with fura-2 were recorded from a fluorescence microscope. Transients stimulated by membrane depolarization (suction electrode or elevated [K+]o) or 15 mM caffeine to release Ca2+ from the sarcoplasmic reticulum (SR) were analyzed for net amplitude, maximal rate of rise (VR), average rate of decline (VR) in [Ca2+]i, and duration. RESULTS: Enflurane and isoflurane reduced electrically stimulated Ca2+i transients in a dose-dependent manner. Enflurane depressed the Ca2+i transient amplitude more than isoflurane. Enflurane was more effective than isoflurane in reducing VR and VF in a concentration-dependent manner. At similar concentrations, both enflurane and isoflurane reduced the steady state elevation of [Ca2+]i by 50 mM K+o. Similarly, enflurane and isoflurane depressed caffeine-sensitive release of Ca2+ from the SR. The reduction in the Ca2+i transient because of SR Ca2+ release was greater in enflurane than in equal concentrations of isoflurane. Rates of elevation and decline in [Ca2+]i were also reduced in enflurane and isoflurane. CONCLUSIONS: The negative inotropic actions of enflurane and isoflurane involve a depression of Ca2+ influx during membrane excitation, as well as a reduction in SR Ca2+ release. Slowed rates of elevation in [Ca2+]i indicate that the latter mechanism may, in part, be caused by alterations in the kinetics of SR Ca2+ release.

Halothane alters control of intracellular Ca2+ mobilization in single rat ventricular myocytes.

In an attempt to understand the cellular mechanisms underlying volatile anesthetic-induced myocardial depression, halothane-induced negative inotropy was investigated in an animal model through continuous monitoring of intracellular Ca2+ concentration [( Ca2+]i) in rat ventricular myocytes loaded with fura-2. Single cells were stimulated with 15 mM caffeine or 15 mM extracellular K+ (K+O) or were paced by extracellular glass suction pipette electrode. With each stimulus modality, halothane (0.6-1.5%) caused a significant (P less than 0.05) and dose-dependent depression of the Ca2+ transient. Caffeine and electrically stimulated Ca2+ transients were reduced, in 1.5% halothane, to 35 +/- 14 and 42 +/- 8% of control, respectively. Resting or basal [Ca2+]i was unaffected by halothane. Halothane did not elicit spontaneous Ca2+ transients in these cells. Single cells stimulated by trains of electrical stimuli at 1.0, 1.5, and 2.0 Hz showed a change in [Ca2+]i from prestimulus levels to a stimulated baseline steady state that appeared to increase with stimulus frequency. Halothane at 0.7% increased the change in resting to stimulated baseline [Ca2+]i and depressed net transients (P less than 0.05) at 1.0 and 1.5 Hz. In contrast, 0.1 microM ryanodine depressed the Ca2+ transients in myocytes stimulated by trains of stimuli, but did not potentiate the change in stimulated baseline [Ca2+]i at any pacing rate. The results are consistent with the hypothesis that halothane reduces Ca2+i availability by causing a net loss of Ca2+ from the sarcoplasmic reticulum. The results from experiments using onset of pacing to induce a sudden increase in Ca2+i load in previously quiescent myocytes suggest that halothane may act to limit sarcoplasmic reticulum and/or sarcolemmal uptake/extrusion mechanisms, as compared to ryanodine, which depletes sarcoplasmic reticulum Ca2+ stores without affecting reuptake and extrusion.

Outward potassium currents in freshly isolated smooth muscle cell of dog coronary arteries.

Outward membrane currents were characterized in single coronary smooth muscle cells of adult beagle dogs. The cells averaged 96.4 x 7.1 microns and had a resting potential of -50.7 mV, an input resistance of 307.9 M omega, a capacitance of 32.3 pF, and a calculated membrane surface area of 4,037 microns2. The cells contracted in response to external application of acetylcholine or high K+. In voltage clamp by use of the suction pipette method, outward current began to appear at -50 mV and reached 15.2 nA at 50 mV with a current density of 376.5 microA/cm2. The current was reduced by external tetraethylammonium, Ba2+, and internal Cs+, and its reversal potential had a Nernst relation to external K+ concentration. Elevation of external Ca2+ (Ca2+o) from 0 to 0.3 mM increased total K+ current by up to 300%; elevation of internal Ca2+ (Ca2+i) to 5 x 10(-7) M by internal perfusion increased total outward current to a similar extent, suggesting a large difference in Ca2+ transmembrane sensitivity. Total whole-cell K+ current consisted of two components: an initial time-independent current (Ii) followed by a time-dependent current (It). Ii and It were through separate K+ channels based on differences in a) sensitivity to Ca2+09b) modulation by an inward Ca2+ current, c) current amplitudes and activation kinetics, and d) responses to pharmacological agents. It was the largest component, measuring 4.5 nA in 0 mM Ca2+o but increasing to 11.9 nA in 0.3 mM Ca2+o with a steep 2.5 power function. It activated with a biexponential time course; in Ca2+o-free solution, its time course was relatively insensitive to voltage changes but became voltage sensitive in the presence of Ca2+o. Further, such sensitivity was abolished or enhanced by Co2+ or Bay K 8644, respectively. We concluded that there are two types of Ca2+-sensitive K+ currents, Ii and It, in coronary smooth muscle cells. Via an inward Ca2+ channel Ca2+o strongly modulates It, both in amplitude and kinetics.

Electrophysiological mechanisms of minoxidil sulfate-induced vasodilation of rabbit portal vein.

The electrophysiological and mechanical properties of the vasodilator minoxidil sulfate (MNXS) were examined in isolated smooth muscle cells and strips from rabbit portal vein. At micromolar concentrations, MNXS inhibited norepinephrine (0.1-1.0 microM)-induced contractions in isolated muscle strips. In isolated cells, norepinephrine caused a dose-dependent depolarization of the resting membrane potential, which was significantly attenuated by MNXS (5 microM); MNXS alone caused a hyperpolarization of the membrane potential. This hyperpolarization was insensitive to Na+-K+ pump blockade by ouabain, but was inhibited by the K+ channel antagonist, tetraethylammonium (20 mM). In voltage-clamp experiments, a resting (background) conductance associated with the resting membrane potential was identified. This conductance, which previously has been shown to be reduced by Ba2+ as well as tetraethylammonium, was increased by MNXS (2 microM). In additional experiments, whole-cell L-type Ca2+ currents were inhibited by micromolar concentrations of MNXS. These experiments show that concentrations of MNXS that inhibit norepinephrine-induced contractions promote K+ conductance and inhibit Ca2+ entry through voltage-dependent Ca2+ channels in vascular smooth muscle cells. These electrophysiological effects of MNXS may be responsible for the vasorelaxant effects of the drug observed in vitro and in vivo.

Effects of tetraethylammonium and 4-aminopyridine on the plateau potential of circular myometrium from the pregnant rat.

In the pregnant rat, spontaneous electrical activity of circular muscle (CM) changes from single, plateau-type action potentials at early and mid-term to repetitive spike trains at term. To examine mechanisms underlying the plateau, we studied the effects of potassium channel blockers tetraethylammonium (TEA) and 4-aminopyridine (4-AP) on membrane potentials in CM from rats on gestation Days 14, 15, 16, 21 (term). Apparent membrane conductance was measured at rest and during the plateau in Day 14 muscles with and without TEA. 4-AP depolarized the resting membrane on all gestation days. Therefore, a direct action of 4-AP on plateau configuration could not be separated from an indirect effect of depolarization. TEA did not affect the resting potential but increased action potential size and depolarization rate on all gestation days. On Day 16, TEA reduced plateau amplitude, unmasking small, repetitive depolarizations. D-600 decreased plateau amplitude and duration and attenuated these effects of TEA. Plateau conductance increased initially then decreased before membrane repolarization. Membrane conductance and outward rectification during the plateau were reduced by TEA. The plateau potential may result from an outwardly rectifying TEA-sensitive current combined with a slow inward current, the plateau magnitude being determined by the relative intensity of each current.

Effect of angiotensin II on vascular resistance in whole-body perfused dogfish.

1. The effect of angiotensin II (AII), norepinephrine (NE), epinephrine (E) and isoproterenol (ISO) was observed on the branchial and systemic circulations in a whole-body-pump perfused dogfish preparation. 2. NE and E increased systemic blood flow resistance, but decreased branchial resistance. 3. ISO decreased both systematic and branchial blood flow resistance. 4. AII had no significant effect on either branchial or systemic resistance.

Vascular volume-distensibility characteristics of the isolated dogfish gut.

The vascular capacitance and volume distensibility of the isolated dogfish gut and segments of dogfish arteries and veins were investigated. The volume-distensibility curves for dogfish arteries and veins are very similar to comparable curves derived from arteries and veins of dogs or man. The vascular volume-distensibility curve of the gut, however, shows a greater distensibility at higher systemic pressure than at lower pressure. Evidence is presented that significant amounts of fluid leave the vascular compartment at a lower systemic pressure than in an isolated dog hindlimb preparation. However, this alone does not explain the atypical vascular volume-distensibility curve obtained from the dogfish gut. It is suggested that in the dogfish capillary filtration pore enlargement takes place at a very low capillary pressure or volume (compared to mammals) and this complicates the construction of a volume-distensibility curve because initial vascular volume is not constant.