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N J Rusch

Publications and source records attributed to N J Rusch.

At least 19 recordsLinked to original sources
PubMed2001

High glucose impairs voltage-gated K(+) channel current in rat small coronary arteries.

Hyperglycemia is associated with impaired endothelium-dependent dilation that is due to quenching of NO by superoxide (O(2)(. -)). In small coronary arteries (CAs), dilation depends more on smooth muscle hyperpolarization, such as that mediated by voltage-gated K(+) (Kv) channels. We determined whether high glucose enhances O(2)(.-) production and reduces microvascular Kv channel current and functional responses. CAs from Sprague-Dawley rats were incubated 24 hours in medium containing either normal glucose (NG, 5.5 mmol/L D-glucose), high glucose (HG, 23 mmol/L D-glucose), or L-glucose (LG, 5.5 mmol/L D-glucose and 17 mmol/L L-glucose). O(2)(.-) production was increased in HG arteries. Whole-cell patch clamping showed a reduction of 4-aminopyridine (4-AP)-sensitive current (Kv current) from smooth muscle cells of HG CAs versus NG CAs or versus LG CAs (peak density was 9.95+/-5.3 pA/pF for HG versus 27.8+/-6.8 pA/pF for NG and 28.5+/-5.2 pA/pF for LG; P<0.05). O(2)(.-) generation (xanthine+xanthine oxidase) decreased K(+) current density, with no further reduction by 4-AP. Partial restoration was observed with superoxide dismutase and catalase. Constriction to 3 mmol/L 4-AP was reduced in vessels exposed to HG (13+/-5%, P<0.05) versus NG (30+/-7%) or LG (34+/-4%). Responses to KCl and nifedipine were not different among groups. Superoxide dismutase and catalase increased contraction to 4-AP in HG CAs. This is the first direct evidence that exposure of CAs to HG impairs Kv channel activity. We speculate that this O(2)(.-)-induced impairment may reduce vasodilator responsiveness in the coronary circulation of subjects with coronary disease or its risk factors.

4-Aminopyridine↗
PubMed2001

Down-regulation of L-type calcium channels in inflamed circular smooth muscle cells of the canine colon.

BACKGROUND & AIMS: Circular smooth muscle phasic contractions and tone are suppressed during colonic inflammation, but the contributing factors are poorly understood. This study investigated if the expression level of voltage-gated long-lasting (L-type) Ca(2+) channel protein and functional Ca(2+) channel current are down-regulated in the circular muscle cells of the inflamed canine colon. METHODS: L-type Ca(2+) channel expression was compared between normal and inflamed smooth muscle cells by Western immunoblots using an antibody directed against the pore-forming alpha 1C-subunit, and patch-clamp methods were used to evaluate Ca(2+) channel current density. RESULTS: The expression of the L-type Ca(2+) channel protein was significantly reduced in inflamed compared with normal circular smooth muscle cell membranes, and this finding was associated with suppressed levels of Ca(2+) channel current in patch-clamped cells. The L-type Ca(2+) channel current in normal and inflamed cells increased proportionately in response to Bay K 8644, but the maximal current density was still lower in the inflamed cells. Acetylcholine increased the L-type Ca(2+) channel current in normal but not in inflamed cells. CONCLUSIONS: The expression level of L-type Ca(2+) channels is down-regulated in the circular smooth muscle cell membranes of the inflamed colon, which may result in reduced Ca(2+) influx. The functional and pharmacologic properties of the channels seem normal. Although some Ca(2+) channels are still present in the inflamed cells, acetylcholine does not activate these channels, which may be caused by additional upstream defects in the receptor signaling cascade. The down-regulation of L-type Ca(2+) channel expression may suppress circular smooth muscle contractions in the inflamed colon and contribute to the abnormalities in motility and digestion observed during inflammatory disorders.

3-Pyridinecarboxylic acid, 1,4-dihydro-2,6-dimethy↗
PubMed1999

Electrical and mechanical responses of rat middle cerebral arteries to reduced PO2 and prostacyclin.

Isolated rat middle cerebral arteries were perfused and superfused with physiological salt solution equilibrated with a control (approximately 140 mmHg) or reduced (approximately 35-40 mmHg) PO2. In other experiments, cerebral arteries were isolated and prostacyclin release was determined by radioimmunoassay for 6-ketoprostaglandin F1alpha. Equilibration of the vessels with reduced PO2 (35 mmHg) solution caused a significant increase in prostacyclin release relative to control PO2 (140 mmHg) conditions. Exposure of middle cerebral arteries to reduced PO2 caused vascular smooth muscle (VSM) hyperpolarization and vessel relaxation, which could be blocked by 1 microM glibenclamide, an inhibitor of the ATP-sensitive K+ channel, but not by 1 mM tetraethylammonium (TEA), an inhibitor of the Ca2+-activated K+ channel. Glibenclamide also inhibited VSM hyperpolarization and vasodilation in response to the stable prostacyclin analog iloprost, but TEA did not affect iloprost-induced dilation of the vessel. Endothelial removal eliminated the electrical and mechanical responses of the arteries to reduced PO2, but vessel responses to iloprost were similar to those of intact vessels. The results of this study are consistent with the hypothesis that hypoxic dilation of rat middle cerebral arteries is due to VSM hyperpolarization mediated by prostacyclin-induced activation of glibenclamide-sensitive K+ channels.

Animals↗
PubMed1999

Loss of endothelium and receptor-mediated dilation in pial arterioles of rats fed a short-term high salt diet.

A high salt diet often is regarded as an accessory risk factor in hypertension, coincidental to the deleterious effect of high blood pressure on vasodilator function. The aim of this study was to determine whether short-term ingestion of a high salt diet per se impairs vasodilator function in the cerebral circulation independent of blood pressure changes. Adult Sprague-Dawley rats were fed a normal salt (0.8%) or high salt (4%) diet for 3 days. Mean arterial pressures were similar in the normal and high salt groups (123+/-2 and 125+/-2 mm Hg, respectively). Subsequently, the responses of the in situ pial arterioles to acetylcholine, iloprost, and sodium nitroprusside were determined in cranial windows using intravital videomicroscopy. Pial arterioles of rats fed normal and high salt diets showed similar resting diameters of 69+/-2 and 72+/-3 microm, respectively, but their reactivity patterns to vasodilator stimuli were markedly different. Arterioles of rats fed a normal salt diet dilated progressively up to 17+/-3% in response to the endothelium-dependent agent acetylcholine (10(-9) to 10(-6) mol/L) and dilated by 22+/-2% in response to the prostaglandin I2 receptor agonist iloprost (3x10(-11) mol/L). In contrast, pial arterioles of rats fed a high salt diet constricted by 4+/-3% and 8+/-2% in response to acetylcholine and iloprost, respectively. Sodium nitroprusside (10(-6) mol/L), a nitric oxide donor, dilated pial arterioles of rats fed low and high salt diets by a similar amount (19+/-3% and 16+/-2%, respectively), suggesting that signaling mechanisms for dilation distal to the vascular smooth muscle membrane were intact after high salt intake. These results provide the first evidence that the short-term ingestion of a high salt diet may severely impair the vasodilator function of the in situ cerebral microcirculation independent of blood pressure elevation.

Animals↗
PubMed1998

Increased expression of Ca2+-sensitive K+ channels in the cerebral microcirculation of genetically hypertensive rats: evidence for their protection against cerebral vasospasm.

The Ca2+-sensitive K+ channel (K(Ca) channel) plays a key role in buffering pressure-induced constriction of small cerebral arteries. An amplified current through this channel has been reported in vascular smooth muscle cells obtained from hypertensive animals, implying that the expression or properties of K(Ca) channels may be regulated by in vivo blood pressure levels. In this study, we investigated this hypothesis and its functional relevance by comparing the properties, expression levels, and physiological role of K(Ca) channels in cerebral resistance arteries from normotensive and genetically hypertensive rats. Whole-cell patch-clamp experiments revealed a 4.7-fold higher density of iberiotoxin-sensitive K(Ca) channel current at physiological membrane potentials in spontaneously hypertensive rat (SHR) compared with Wistar-Kyoto (WKY) rat cerebrovascular smooth muscle cells (n = 18 and 21, respectively). However, additional single-channel analysis in detached patches showed similar levels of unitary conductance, voltage, and Ca2+ sensitivity in K(Ca) channels from WKY and from SHR membranes. In contrast, Western analysis using an antibody directed against the K(Ca) channel alpha-subunit revealed a 4.1-fold increase in the corresponding 125-kD immunoreactive signal in cerebrovascular membranes from SHR compared with WKY rats. The functional impact of this enhanced K(Ca) channel expression was assessed in SHR and WKY rat pial arterioles, which were monitored by intravital microscopy through in situ cranial windows. Progressive pharmacological block of K(Ca) channels by iberiotoxin (0.1 to 100 nmol/L) dose-dependently constricted pial arterioles from SHR and WKY rats (n = 6 to 8). The arterioles in SHR constricted 2- to 4-fold more intensely, and vasospasm occurred in some vessels. These data provide the first direct evidence that elevated levels of in situ blood pressure induce K(Ca) channel expression in cerebrovascular smooth muscle membranes. This homeostatic mechanism may critically regulate the resting tone of cerebral arterioles during chronic hypertension. Furthermore, the overexpression of distinct K+ channel types during specific cardiovascular pathologies may provide for the upregulation of novel disease-specific membrane targets for vasodilator therapies.

Analysis of Variance↗
PubMed1998

Intracellular acidosis differentially regulates KV channels in coronary and pulmonary vascular muscle.

Decreases in intracellular pH (pHi) potently dilate coronary resistance arteries but constrict small pulmonary arteries. To define the ionic mechanisms of these responses, this study investigated whether acute decreases in pHi differentially regulate K+ currents in single vascular smooth muscle (VSM) cells isolated from rat coronary and pulmonary resistance arteries. In patch-clamp studies, whole cell K+ currents were elicited by 10-mV depolarizing steps between -60 and 0 mV in VSM cells obtained from 50- to 150-micrometers-OD arterial branches, and pHi was lowered by altering the NH4Cl gradient across the cell membrane. Progressively lowering pHi from calculated values of 7.0 to 6.7 and 6.4 increased the peak amplitude of K+ current in coronary VSM cells by 15 +/- 5 and 23 +/- 3% but reduced K+ current in pulmonary VSM cells by 18 +/- 3 and 21 +/- 3%, respectively. These changes were reversed by returning cells to the control pHi of 7.0 and were eliminated by dialyzing cells with pipette solution containing 50 mmol/l HEPES to buffer NH4Cl-induced changes in pHi. Pharmacological block of ATP-sensitive K+ channels and Ca2+-activated K+ channels by 1 micromol/l glibenclamide and 100 nmol/l iberiotoxin, respectively, did not prevent changes in K+ current levels induced by acidotic pHi. However, block of voltage-gated K+ channels by 3 mmol/l 4-aminopyridine abolished acidosis-induced changes in K+ current amplitudes in both VSM cell types. Interestingly, alpha-dendrotoxin (100 nmol/l), which blocks only select subtypes of voltage-gated K+ channels, abolished the acidosis-induced decrease in K+ current in pulmonary VSM cells but did not affect the acidosis-induced increase in K+ current observed in coronary VSM cells. These findings suggest that opposing, tissue-specific effects of pHi on distinct subtypes of voltage-gated K+ channels in coronary and pulmonary VSM membranes may differentially regulate vascular reactivity in these two circulations under conditions of acidotic stress.

4-Aminopyridine↗
PubMed1998

Distinct endothelial impairment in coronary microvessels from hypertensive Dahl rats.

Hypertension has been linked to an impaired dilator function of the coronary microvascular endothelium in vivo. However, the profile and mechanism of this dysfunction remain obscure. Thus, this study compared diameter responses to acetylcholine (ACH), bradykinin (BKN), and substance P (SP) between coronary microvessels (i.d.=106+/-4 microm) dissected from left ventricles of normotensive and hypertensive Dahl rats (Dahl-NT and Dahl-HT, respectively). Vessels were cannulated and pressurized on glass pipettes at 80 mm Hg, and internal diameters were monitored by videomicroscopy. Coronary microvessels from Dahl-NT and Dahl-HT showed similar dilator responses to ACH (100 pmol/L to 10 micromol/L), with maximal diameter increases of 63+/-5 microm and 63+/-7 microm, respectively (n=31,17). However, only vessels from Dahl-NT showed dilator responses to SP (10 fmol/L to 1 nmol/L) and BKN (100 fmol/L to 10 nmol/L). All dilator responses persisted after N-nitro-L-arginine (10 micromol/L) or indomethacin (10 micromol/L), but were blunted after inhibition of cytochrome P450 by 10 micromol/L octadecynoic acid (n=6-8). These results suggest that: (1) coronary microvessels from Dahl-HT show a unique pattern of endothelial impairment, whereby ACH-induced relaxations persist at a time when dilator responses to SP and BKN are severely blunted, and (2) a cytochrome P450 product, rather than nitric oxide or prostacyclin, may partly mediate the vasodilator responses to ACH, SP and BKN.

Acetylcholine↗
PubMed1997

Hypoxia-induced hyperpolarization is not associated with vasodilation of bovine coronary resistance arteries.

The effect of reduced PO2 on the transmembrane potential and diameter of small cannulated coronary resistance arteries was evaluated by microelectrode and videomicroscopic methods. Bovine coronary resistance arteries (158 +/- 8 microm ID) were cannulated with glass micropipettes and perfused and superfused with physiological salt solution. Lowering the PO2 of the physiological salt solution from 140 +/- 4 to 36 +/- 2 mmHg increased the smooth muscle cell transmembrane potential from -51 +/- 2 to -62 +/- 2 mV in both endothelium-intact and -denuded coronary resistance arteries. This hyperpolarization was blocked by superfusion with the K+-channel blocker glibenclamide (1 microM). However, low PO2 did not significantly dilate either endothelium-intact or -denuded coronary resistance arteries, although superfusion with 1 microM cromakalim, a K+-channel activator, induced a 6-mV hyperpolarization and increased the diameter by 33 +/- 10 microm. These results suggest that reduced PO2 directly hyperpolarizes the vascular smooth muscle of coronary resistance arteries by activation of glibenclamide-sensitive K+ channels, but other nonvascular mechanisms may mediate the vasodilation response to low PO2.

Acetylcholine↗
PubMed1997

Increased expression of Ca2+-sensitive K+ channels in aorta of hypertensive rats.

Potassium efflux through Ca2+-sensitive K+ channels (K[Ca] channels) is increased in arterial smooth muscle cells from hypertensive rats, but the molecular mechanism is unknown. The goal of this study was to compare the levels of K(Ca) channel current between aortic smooth muscle cells from adult Wistar-Kyoto rats (WKY) and spontaneously hypertensive rats (SHR) and then use Western blot methods and ribonuclease protection assays to examine the expression and mRNA levels for the K(Ca) channel in these same vascular tissues. Whole-cell patch-clamp methods indicated a larger component of K(Ca) channel current, sensitive to block by iberiotoxin (100 nmol/L), in single aortic smooth muscle cells from SHR compared with WKY. Subsequent Western blot analysis using a site-specific antibody (anti-alpha[913-926]) directed against the S9/S10 linker of the alpha-subunit of the K(Ca), channel revealed a 125-kD immunoreactive band in lanes loaded with either WKY or SHR aortic muscle membranes. The immunoreactive density of this band, which corresponded to the known molecular size of the alpha-subunit, was 2.2-fold greater in lanes loaded with aortic smooth muscle membranes from the hypertensive animals. However, despite this evidence for an increased expression and functional enhancement of K(Ca) channels in aortic smooth muscle membranes of SHR, ribonuclease protection assays with a 32P-labeled riboprobe targeted against the S9/S10 linker of the K(Ca) channel alpha-subunit revealed no difference in mRNA levels for the alpha-subunit between WKY and SHR aortic tissue. These findings provide initial evidence that (1) an increased expression of K(Ca) channels may be a mechanism for the enhanced K(Ca) current in aortic smooth muscle membranes of SHR, and (2) the upregulation of K(Ca) channels in arterial muscle membranes during hypertension, which is regarded as a homeostatic mechanism for buffering vascular excitability, may rely on posttranscriptional events.

Animals↗
PubMed1997

Enzymatic isolation and characterization of single vascular smooth muscle cells from cremasteric arterioles.

OBJECTIVE: The goal of the present study was to develop a method to isolate viable arteriolar muscle cells from single cremasteric arterioles, which retain the contractile and electrophysiological phenotype of the donor microvessels. METHODS: Arterioles were hand-dissected from rat and hamster cremaster muscles and dissociated by incubation in papain and dithioerythritol for 35 min followed by incubation in collagenase, elastase, and soybean trypsin inhibitor for 10 to 25 min in solutions containing 100 microM Ca2+. 10 microM sodium nitroprusside, and 1 mg/ml albumin at 37 degrees C. RESULTS: Populations of single smooth muscle cells enzymatically isolated from cremasteric arterioles showed elongated fusiform morphology and intact plasmalemmal membranes as indicated by retention of calcein, by exclusion of ethidium homodimer-1 and by high membrane resistances (11 +/- 0.8 C omega, n = 36 for rat cells; 8 +/- 0.6 C omega, n = 21 for hamster cells: p < 0.05). Muscle cells contracted in a concentration-dependent fashion in response to pipette application of norepinephrine (10 nM-100 microM). Cell shortening in response to 1 microM norepinephrine was inhibited by 10 microM phentolamine, 1 microM sodium nitroprusside, and 1 microM nifedipine or nominally Ca(2+)-free media. Resting membrane potential recorded in patch-clamped cells by perforated patch methods was -48 +/- 1 mV (n = 47) for rat cells and -44 +/- 2.8 mV (n = 14) for hamster cells (p > 0.05). Families of voltage-dependent K+ currents were observed during stepwise depolarizing pulses from -60 mV to more positive potentials. Blockers of voltage-gated and ATP-sensitive K+ channels (4-aminopyridine [3 mM] and glibenclamide [1 microM], respectively) inhibited membrane K+ conductance, increased membrane resistance, and depolarized cells by 20 +/- 4 mV (n = 8) and 14 +/- 3 mV (n = 6), respectively. CONCLUSIONS: The present method permits isolation of smooth muscle cells from a single cremasteric arteriole. These cells seem to retain the contractile phenotype, alpha-adrenergic signaling cascade, membrane potential, and K+ conductances described for the donor arteriole. Correlating the functional and electrophysiological properties of these smooth muscle cells to in situ and in vitro studies of their donor arterioles should provide a useful extension for understanding the physiology, pathophysiology, biophysics, and cell biology of the microcirculation in skeletal muscle.

Animals↗
PubMed1996

Endothelium and vascular smooth muscle function in internal mammary artery after cryopreservation.

An optimal cryopreserved arterial conduit should have anatomic and physiologic characteristics similar to those of the fresh artery. We have cryopreserved canine internal mammary artery (IMA) with intact, prostacyclin (PGI2)-producing endothelial cells, but the underlying vascular smooth muscle appeared nonfunctional. Thus the aim of this study was to evaluate which steps of the cryopreservation method compromise vascular smooth muscle function in cryopreserved IMA. Isometric tension recording responses to vasoconstrictor agonists were used to evaluate the level of vascular muscle integrity, while endothelial function was assessed by relaxation responses to acetylcholine and by PGI2 production. These variables were measured in vessels rewarmed to 37 degrees C, following initial exposure to different stages of an eight-step computerized cryopreservation protocol (n = 10). In this protocol, tissue temperature was lowered to -10 degrees C (steps 1-2), rapidly reduced to -12 degrees C (steps 3-4), reduced further to -40 degrees C at a rate of -0.5 degrees C/min (steps 5-6), lowered to -70 degrees C (step 7), and finally equilibrated at - 196 degrees C by immersion in liquid nitrogen (step 8). Compared to fresh IMA, the amplitude of dose-dependent contractions to norepinephrine (NE 10(-9) to 10(-4) M) was step-wise depressed after cooling to < or = -12 degrees C (steps 3-4), showing only 75 and 19% of maximal contraction after steps 3-4 and step 7, respectively. Similarly, depolarization-induced contractions to KCl (10 to 60 mM) also were progressively depressed after step-wise cryopreservation, showing reduced contractile amplitudes even after steps 1-2 (-10 degrees C). Vascular muscle agonist sensitivity was unchanged (KCl) or only mildly influenced (NE) by these same temperature reductions. In contrast, endothelial-dependent relaxation to acetylcholine and PGI2 production were maintained after all steps in the cryopreservation process, and baseline PGI2 production was higher in cryopreserved IMA. These data indicate that IMA smooth muscle cell viability is poorly preserved after cooling to below -10 to -12 degrees C of the cryopreservation process, whereas endothelial cell function appears intact after profound cooling to -196 degrees C. The loss of vascular smooth muscle responsiveness coupled with the protection afforded by an intact endothelium may provide an arterial conduit less susceptible to vasospasm. Such a graft, however, would lack the dynamic properties of flow regulation in response to the metabolic needs of the myocardium.

Acetylcholine↗
PubMed1996

Single-channel and functional characteristics of a KCa channel in vascular muscle membranes of human saphenous veins.

The saphenous vein is used extensively to test for the effects of vasodilator substances on venous reactivity, but the K+ channel types that mediate vasodilation have not been identified. Thus the goal of this study was to identify K+ channel types in vascular smooth muscle membranes of human saphenous vein (HSV), which may contribute to membrane repolarization and control of venous tone. Fourteen HSVs obtained from bypass surgery were enzymatically dissociated into single vascular myocytes for patch-clamp analysis of inside-out patches (n = 81). HSV membranes showed primarily high-conductance (226 pS) K+ channels, which accounted for > or = 95% of total patch current at physiologic voltages. Channels were highly K+ selective, showed steep voltage and Ca2+ sensitivity, and were blocked by 100 nM iberiotoxin and < or = 1 mM tetraethylammonium (TEA). These Ca(2+)-sensitive channels (KCa) also showed stacked openings in depolarized patches exposed to 300-1,000 nM calcium, suggesting multiple functional KCa channels in a single membrane patch. In tension-recording studies, isolated segments of HSV exposed to 100 nM norepinephrine contracted further during progressive block of KCa channels by 0.1-3 mM TEA, suggesting that KCa channels are pathways for repolarization and vasodilation in HSV smooth muscle cells. Our finding of KCa channels in smooth muscle membranes of HSV, if extended to the plasma membranes of other human peripheral veins, suggests that this channel may represent a therapeutic site for alleviation of conditions of increased venous tone.

Adult↗
PubMed1996

Mechanisms for regulation of arterial tone by Ca2+-dependent K+ channels in hypertension.

1. The membrane potential and reactivity of arterial smooth muscle cells is regulated by a variety of K+ channels, which are highly expressed in vascular smooth muscle membranes. 2. Of these K+ channel types, the high-conductance, Ca2+-dependent K+ channel appears to be up-regulated in arterial smooth muscle membranes from hypertensive animals. 3. Patch-clamp studies show that whole-cell membranes and membrane patches of arterial smooth muscle obtained from rats with genetic or renal hypertension show an increased macroscopic and single-channel Ca2+-activated K+ current. Pharmacological block of this K+ current profoundly constricts aortic, renal, mesenteric and femoral arteries obtained from the same hypertensive animals, suggesting that Ca2+-dependent K+ current is a critical determinant of resting membrane potential in arterial muscle exposed to elevated blood pressure. 4. Thus, K+ efflux through Ca2+-dependent K+ channels appears to constitute an important homeostatic mechanism for buffering increases in arterial reactivity in hypertension.

Animals↗
PubMed1996

Enzymatic isolation and characterization of single vascular smooth muscle cells from cremasteric arterioles.

OBJECTIVE: The goal of the present study was to develop a method to isolate enzymatically viable arteriolar muscle cells from single cremasteric arterioles, which retain the contractile and electrophysiological phenotype of the donor microvessels. METHODS: Arterioles were hand-dissected from rat and hamster cremaster muscles and dissociated by incubation in papain and dithioerythritol for 35 min followed by incubation in collagenase, elastase, and soybean trypsin inhibitor for 10 to 25 min in solutions containing 100 microM Ca2+, 10 microM sodium nitroprusside, and 1 mg/ml albumin at 37 degrees C. RESULTS: Populations of single smooth muscle cells enzymatically isolated from cremasteric arterioles showed elongated fusiform morphology and intact plasmalemmal membranes as indicated by retention of calcein, by exclusion of ethidium homodimer-1, and by high membrane resistances (11 +/- 0.8 G omega, n = 36 for rat cells; 8 +/- 0.6 G omega, n = 21 for hamster cells; p < 0.05). Muscle cells contracted in a concentration-dependent fashion in response to pipette application of norepinephrine (10 nM-100 microM). Cell shortening in response to 1 microM norepinephrine was inhibited by 10 microM phentolamine, 1 microM sodium nitroprusside, and 1 microM nifedipine or nominally Ca(2+)-free media. Resting membrane potential recorded in patch-clamped cells by perforated patch methods was -48 +/- 1 mV (n = 47) for rat cells and -44 +/- 2.8 mV (n = 14) for hamster cells (p > 0.05). Families of voltage-dependent K+ currents were observed during stepwise depolarizing pulses from -60 mV to more positive potentials. Blockers of voltage-gated and ATP-sensitive K+ channels (4-Aminopyridine [3 mM] and glibenclamide [1 microM], respectively) inhibited membrane K+ conductance, increased membrane resistance, and depolarized cells by 20 +/- 4 mV (n = 8) and 14 +/- 3 mV (n = 6), respectively. CONCLUSIONS: The present method permits isolation of smooth muscle cells from a single cremasteric arteriole. These cells seem to retain the contractile phenotype, alpha-adrenergic signaling cascade, membrane potential, and K+ conductances described for the donor arteriole. Correlating the functional and electrophysiological properties of these smooth muscle cells to in situ and in vitro studies of their donor arterioles should provide a useful extension for understanding the physiology, pathophysiology, biophysics, and cell biology of the microcirculation in skeletal muscle.

Abdominal Muscles↗
PubMed1996

Potassium channels: new targets for drug treatment.

Potassium channels in cell membranes regulate cellular excitability, proliferation and growth. The central role of these ion channels in cell function has made them the targets for current drug treatment of hypertension, hair loss and diabetes, and also the subject of intense research for new drug therapies.

Diabetes Mellitus↗
PubMed1995

Reactivity of human saphenous veins at arterial perfusion pressures.

Vasospasm of human saphenous vein grafts has been reported after aorta-coronary bypass operations. However, it is unknown whether veno-arterial grafts are inherently responsive to vasoconstrictor stimuli after implantation into the arterial circulation or whether their vasomotion is secondary to hemodynamic changes. Thus in this study we used in vitro methods to directly evaluate whether isolated human saphenous vein segments respond to vasoconstrictor agents at arterial pressure levels. External diameter and intraluminal flow were monitored in 12 human saphenous vein segments, which were perfused at 30 ml/min with physiologic salt solution at 90, 70, and 50 mm Hg. Increasing intraluminal pressure higher than 50 mm Hg or exposing the vein to Ca(2+)-free media did not increase vessel external diameter or intraluminal flow, which suggests that human saphenous veins were fully distended at pressures of 50 mm Hg or greater. However, all human saphenous veins were activated by a 1 mumol/L dose of norepinephrine at 50 mm Hg and dilated during subsequent intraluminal infusion of a 1 mumol/L dose of acetylcholine, showing intact vascular smooth muscle and endothelial cell function. In the same vessels, a 1 mumol/L concentration of 5-hydroxytryptamine constricted human saphenous veins by 19%, 22%, and 26% at intraluminal pressures of 90, 70, and 50 mm Hg, respectively, and reduced vessel flow by 6%, 24%, and 42% at the same pressure levels. Similarly, a 1 mumol/L concentration of norepinephrine constricted vessels pressurized at 90, 70, and 50 mm Hg by 9%, 12%, and 17%, respectively, and attenuated vessel flow by as much as 32%. We conclude that human saphenous vein segments are fully distended at perfusion pressures greater than 50 mm Hg, but can dynamically constrict to vasoactive agonists and regulate graft flow at intraluminal pressures as high as 90 mm Hg. Our findings in isolated human saphenous vein segments lend support to clinical observations that human saphenous vein grafts should be regarded as vasoactive conduits after implantation at arterial pressure levels.

Acetylcholine↗
PubMed1995

Halothane and isoflurane decrease the open state probability of K+ channels in dog cerebral arterial muscle cells.

BACKGROUND: Both halothane and isoflurane evoke cerebral vasodilation. One of the potential mechanisms for arterial vasodilation is enhanced K+ efflux resulting from an increased opening frequency of membrane K+ channels. The current study was designed to determine the effects of volatile anesthetics on K+ channel current in single vascular smooth muscle cells isolated from dog cerebral arteries. METHODS: Patch clamp recording techniques were used to investigate the effects of volatile anesthetics on macroscopic and microscopic K+ channel currents. RESULTS: In the whole-cell patch-clamp mode, in cells dialyzed with pipette solution containing 2.5 mM EGTA and 1.8 mM CaCl2, depolarizing pulses from -60 to +60 mV elicited an outward K+ current that was blocked 65 +/- 5% by 3 mM tetraethylammonium (TEA). Halothane (0.4 and 0.9 mM) depressed the amplitude of this current by 18 +/- 4% and 34 +/- 6%, respectively. When 10 mM EGTA was used in the pipette solution to strongly buffer intracellular free Ca2+, an outward K+ current insensitive to 3 mM TEA was elicited. This K+ current, which was reduced 51 +/- 4% by 1 mM 4-aminopyridine, was also depressed by 17 +/- 5% and 29 +/- 7% with application of 0.4 and 0.9 mM halothane, respectively. In cell-attached patches using 145 mM KCl in the pipette solution and 5.2 mM KCl in the bath, the unitary conductance of the predominant channel type detected was 99 pS. External application of TEA (0.1 to 3 mM) reduced the unitary current amplitude of the 99 pS K+ channel in a concentration-dependent manner. The open state probability of this 99 pS K+ channel was increased by 1 microM Ca2+ ionophore (A23187). These findings indicate that the 99 pS channel measured in cell-attached patches was a TEA-sensitive, Ca(2+)-activated K+ channel. Halothane and isoflurane reversibly decreased the open state probability (NPo), mean open time, and frequency of opening of this 99 pS K+ channel without affecting single channel amplitude or the slope of the current-voltage relationship. CONCLUSIONS: Halothane and isoflurane suppress the activity of K+ channels in canine cerebral arterial cells. These results suggest that mechanisms other than K+ channel opening likely mediate volatile anesthetic-induced vasodilation.

Animals↗