Regulation of intracellular pH and its role in vascular smooth muscle function.
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Animals↗
Biomedical subjects
C Aalkjaer
Publications and source records attributed to C Aalkjaer.
Mechanisms behind the relaxing effect of furosemide on the isolated rabbit ear artery.
The effect of furosemide on isometric contraction and 86Rb uptake were studied in the isolated rabbit central ear artery (CEA). A concentration-dependent relaxing effect of furosemide (0.06 mM-1.0 mM) was found in vessel segments with intact endothelium. The maximal relaxation was 28.6 +/- 3.9% (10). The effect was not diminished in segments deprived of endothelium, and removal of endothelium itself caused no change of the force development to electrical field stimulation. The relaxing effect was time-dependent and stimulation-dependent and was not significantly affected by membrane depolarization induced by increasing external [K+] from 10 to 120 mM. The 86Rb uptake was inhibited by both furosemide and ouabain (8.0 +/- 0.5(8) and 5.3 +/- 0.5(8) versus 12.8 +/- 0.9(16) nmol (K+).mm-1.(10 min.)-1 in the furosemide (1.0 mM), ouabain (1.0 mM) and control groups, respectively) without interaction between the two drugs. The 86Rb uptake was not further inhibited by increasing the furosemide concentration from 0.12 mM to 1.0 mM. Our results suggest: firstly, the direct relaxing effect of furosemide on isolated vessel segments is endothelium-independent and secondly, the inhibition of the Na(+)-K(+)-Cl- cotransport and a possible consequent hyperpolarization of the membrane is unlikely to be the sole mechanism responsible for the vasorelaxant effect of furosemide. The demonstrated direct effect on vascular tone may be of clinical importance in situations with very high plasma concentrations of the drug or very low concentrations of serum albumin.
Animals↗
Effect of pinacidil on ion permeability in resting and contracted resistance vessels.
Pinacidil is thought to cause vasodilatation by opening K+ channels and consequent hyperpolarization. This proposed mechanism of action is based mainly on membrane potential measurements and 42K or 86Rb efflux experiments under resting conditions. We have measured the simultaneous effect of pinacidil on force and membrane potential in resting and norepinephrine-contracted rat mesenteric resistance vessels. Also the effect of pinacidil on 42K and 36Cl efflux and 22Na uptake in the absence and presence of norepinephrine was examined. From the membrane potential and ion flux measurements the ion permeabilities were calculated. In both resting and norepinephrine-contracted vessels, pinacidil caused a large hyperpolarization, the latter situation being associated with an almost complete relaxation. In both resting and norepinephrine-stimulated vessels, pinacidil caused a large increase in K+ permeability and a decrease in Cl-permeability, whereas no significant change of Na+ permeability was found. Our results suggest that pinacidil causes vasodilation due to hyperpolarization. The major cause for the hyperpolarization is an increase in K+ permeability.
Animals↗
Structure and function of small arteries.
The small arteries (prearteriolar vessels with lumen diameter less than approximately 500 microns) contribute importantly to and participate actively in the regulation of the peripheral resistance. New techniques, building on the classic histological and hemodynamic techniques, have enabled detailed in vitro investigation of small arteries. At present, research in small arteries is in its infancy, and our understanding of the heterogeneity of small arteries within vascular beds, between vascular beds, and between species is extremely limited. This review attempts to describe the current status of the field. New techniques, based primarily on a wire myograph (where the vessels are mounted as ring preparations) and a pressure myograph (where vessels are cannulated and pressure-lumen relations are determined), have allowed in vitro investigations of small arteries. The more physiological arrangement of the pressure myograph allows, for example, investigation of the vasoconstrictor response to raised intravascular pressure (the Bayliss response), whereas the less-sophisticated wire myograph is similar to use and may be more useful in certain situations where particular mechanisms are being investigated. Both techniques allow simultaneous measurements of vessel tone and a variety of parameters (e.g., membrane potential and intracellular ion activities) and thus allow precise determination of the relation between small artery structure and function. The vessels appear to remain fully viable with regard to the contractility of their smooth muscle cells as well as to the function of their perivascular nerves and their endothelium. The evidence suggests that the monovalent transport mechanisms in the plasma membrane, in particular potassium channels, play an important role in the determination of the membrane potential in small arteries, although the relation is more complex than indicated by the Goldman equation. Confirmation of these findings requires, however, simultaneous determinations of ion transport and vascular tone under conditions where vessels are subjected to mechanical loading. The membrane potential, through its effect on potential-dependent calcium channels, plays an important role in the determination of vascular tone. With regard to calcium homeostasis, current knowledge is hampered by the lack of direct measurements of the relation between cytoplasmic calcium and vascular tone. The evidence, however, suggests that besides potential-dependent calcium channels, receptor-operated calcium channels are present in the plasma membrane, although this still requires confirmation. The role of the sarcoplasmic reticulum is not clarified.(ABSTRACT TRUNCATED AT 400 WORDS)
Arteries↗
Mechanism of the vasodilator action of pinacidil.
The mechanism of the vasodilator action of pinacidil has been studied in rat mesenteric small arteries. The results show, first, that the use of flux studies to make measurements of ion permeability requires knowledge of the membrane potential, especially as regards K+ permeability. Second, the results confirm that the vasodilator effect of pinacidil is due to an increase in K+ permeability. Lastly, the results suggest that the K+ channels involved are sensitive to glibenclamide.
Animals↗
The contractile effects of porcine tetradecapeptide renin substrate in human resistance vessels: evidence of activation by vascular wall renin and serine proteases.
In order to test the hypothesis that the contraction induced in human resistance arterioles by porcine tetradecapeptide renin substrate (TDP) is mediated by enzymes specific to the renin-angiotensin cascade, human resistance vessels from skin and subcutaneous fat were mounted in a myograph and exposed to TDP in the presence and absence of the human renin inhibitor H261, the serine protease inhibitor aprotinin, a polyclonal anti-human renin antibody and captopril. TDP induced a dose-dependent contraction that could be abolished by saralasin. The sensitivity to TDP was significantly attenuated by H261, aprotinin and combinations of captopril with aprotinin and captopril plus aprotinin and H261, as indicated by a significant reduction in pD2 (-log10ED50 [mmol/l]) for TDP. However, captopril alone was ineffective. It was concluded that at physiological pH, porcine TDP induces contraction in human resistance vessels by the action of enzymes not specific to the renin-angiotensin cascade. Whilst a clear inhibitory effect of H261 was demonstrated, a significant comparable inhibition by captopril and a polyclonal renin antibody was not observed. This may reflect the difficulty with which these inhibitors gain access to their intracellularly located substrates.
Electric Stimulation↗
Abnormal structure and function of isolated subcutaneous resistance vessels from essential hypertensive patients despite antihypertensive treatment.
The morphological and functional characteristics of isolated subcutaneous resistance vessels (about 230 microns internal diameter) from 13 patients treated for essential hypertension for a median period of 14 months and from 15 matched normotensive controls were examined. The blood pressure of the patients and the controls were not significantly different at the time of examination. However, although compared with the controls, the lumen diameter of the vessels from the patients was not significantly different, the media thickness to lumen diameter ratio was 19% greater. Furthermore, although there was no difference in the active pressure response of the vessels from the two groups, the vessels from the patients had a lower sensitivity to calcium, relaxed faster after a contraction and the sensitivity to exogenous noradrenaline shifted more to the left with cocaine. Since the abnormalities found here have previously also been found in vessels from patients with untreated essential hypertension, the study suggests that despite antihypertensive treatment to normotensive levels for about 1 year, some morphological as well as functional characteristics of the resistance arteries are not fully normalized. This could have consequences for the prognosis of essential hypertension.
Antihypertensive Agents↗
Effects of drug treatment on human resistance arteriole morphology in essential hypertension: direct evidence for structural remodelling of resistance vessels.
To examine whether effective drug treatment would reverse the morphological changes in resistance arterioles which characterise untreated essential hypertension, the vessels' media thickness and contractility were measured before and after long-term treatment. Nine patients underwent skin biopsy before and after a mean 13 months of antihypertensive treatment (range 4-24 months). Subcutaneous resistance arterioles were dissected and mounted in a myograph. Treatment did not affect the shift in sensitivity to noradrenaline in the presence of cocaine, sensitivity to exogenously administered calcium, and the rate of relaxation after withdrawal of vasoconstrictor stimuli. The media/lumen ratio fell significantly (p = 0.011) owing to a significant regression in media thickness (p = 0.0195), but this was not complete. Thus, effective antihypertensive treatment can reverse some of the structural change observed in the untreated state, although it apparently does not affect calcium sensitivity or relaxation rate; these features are thought to be consequences of the disease after it has become established.
Antihypertensive Agents↗
Vasopressin-mediated contractions in mesenteric arteries from spontaneously hypertensive rats: differences in response compared with Wistar-Kyoto animals.
1. The sensitivity of mesenteric resistance arterioles to [arginine]vasopressin (AVP) was investigated in spontaneously hypertensive rats (SHR) and control Wistar-Kyoto rats (WKY). No difference in pAVP (-log dose of AVP producing 50% of the maximum response) was observed [SHR 0.21 +/- 0.03 m-units/ml (n = 10) vs WKY 0.15 +/- 0.06 m-units/ml (n = 9)], although SHR vessels exhibited greater absolute tension development. 2. Both strains of rat displayed tachyphylaxis to repeated stimulation with AVP, and oscillatory tension changes were observed in all vessels from SHR and rarely in WKY vessels at activating concentrations of AVP. 3. AVP did not elicit a contractile response after noradrenaline-induced calcium depletion. 4. After vessels were depleted of calcium by using a combination of calcium-free media and noradrenaline stimulation, restoration of calcium in the presence of AVP elicited a greater contractile response in SHR vessels. 5. The results therefore provide evidence for an increased calcium response to AVP in SHR resistance vessels, although this was only demonstrable by calcium recovery experiments.
Animals↗
Effect of pinacidil on norepinephrine- and potassium-induced contractions and membrane potential in rat and human resistance vessels and in rat aorta.
The effect of pinacidil on contractile responses to norepinephrine, potassium, and membrane potential was examined in rat and human resistance vessels. In some experiments rat aorta was also used. Pinacidil (0.1-30 microM) caused a concentration-dependent relaxation of norepinephrine-induced contractions in all vessels studied. In the same concentration range, pinacidil had only little effect on potassium (125 mM) activated rat mesenteric and femoral resistance vessels. In denervated rat mesenteric resistance vessels, a depolarization with potassium (125 mM) before superimposing a norepinephrine tone markedly diminished the effect of pinacidil. In resting rat mesenteric resistance vessels, pinacidil (1-10 microM) caused a hyperpolarization of 10-15 mV. In rat aorta, pinacidil (10 microM) caused a significant (p less than 0.001) increase in 86Rb+ efflux rate constant whereas 1 microM had no effect. The results of these experiments indicate that the vasodilating effect may be caused by a hyperpolarization of the vascular smooth muscle cell membrane.
Aged↗
Pinacidil opens K+-selective channels causing hyperpolarization and relaxation of noradrenaline contractions in rat mesenteric resistance vessels.
1. The effects of pinacidil on noradrenaline-induced tone, smooth muscle membrane potential and 42K- and 86Rb-efflux from isolated mesenteric resistance vessels (internal diameter 200 microns) of the rat have been studied. 2. Pinacidil (0.3-10 microM) produced concentration-dependent suppression of noradrenaline-induced tone. 3. Pinacidil (0.3-10 microM) caused concentration-dependent hyperpolarization of the smooth muscle. 4. In rat resistance vessels loaded with 42K, pinacidil (1-10 microM) significantly increased the 42K-efflux rate constant. 5. With the use of 86Rb as a marker for K+, 1 microM pinacidil did not affect the 86Rb-efflux rate constant, while 10 microM pinacidil transiently increased the 86Rb rate constant. 6. The results indicate that the relaxant action of pinacidil in these vessels is due to the opening of K+-channels and consequent hyperpolarization. The K+-channels opened are selective for 42K over 86Rb.
Animals↗
Intracellular pH regulation in resting and contracting segments of rat mesenteric resistance vessels.
1. The pH-sensitive dye 2',7'-bis-(2-carboxyethyl)-5 (and -6)-carboxyfluorescein (BCECF) was used to measure intracellular pH (pHi) in segments of rat resistance vessels (internal diameter about 200 microns) with the vessels mounted in a myograph for simultaneous measurements of isometric contraction. 2. BCECF loaded slowly into the vessels over 1 h and did not affect the maximal contractility of the vessels. There was a loss of dye with time which, however, was very slow when the segments were only excited for 2 s/min, suggesting that the loss was mainly due to dye bleaching with only a very slow leak. 3. The ratio of the emissions (at 540 nm) with excitation at 495 and 450 nm was calibrated in terms of pH using the K+-H+ ionophore nigericin. This calibration gave a pHi value of 7.15 +/- 0.02 (n = 20), suggesting that hydrogen ions are not in electrochemical equilibrium in these vascular smooth muscles which have a membrane potential of about -60 mV. 4. Addition of 10 mM-NH4Cl caused a transient alkalinization and wash-out of 10 mM-NH4Cl a transient acidification. Increasing CO2 with maintained bicarbonate caused a rapid acidification followed by an incomplete recovery. Removal of CO2 and bicarbonate (HEPES-buffered solution) with constant extracellular pH caused a transient alkalinization but steady-state pHi was not significantly altered. 5. In bicarbonate-free buffer the Na+-H+ exchange blocker 5-(N-ethyl-N-isopropyl) amiloride (EIPA) and sodium-free conditions caused a slow acidification. In bicarbonate buffer (PSS) EIPA had no detectable effect after 10 min but the anion exchange blocker diisothio-cyanatostilbenedisulphonic acid (DIDS) caused a small acidification over that time course. 6. The rate of recovery after an acid load was about 50% lower in HEPES buffer compared to PSS and it was inhibited by EIPA. In PSS amiloride and EIPA each had a small inhibitory effect on the pH recovery after an acid load. DIDS also inhibited the recovery from an acid load in PSS and this effect was additive to that of EIPA. DIDS and EIPA also had additive inhibitory effects on the 22Na+ influx stimulated by the acid loading, while in HEPES buffer DIDS had no effect on either pH recovery or 22Na+ influx. These results suggest that a Na+-H+ exchange and an influx of bicarbonate coupled to sodium influx are of importance for pHi control in these vessels.(ABSTRACT TRUNCATED AT 400 WORDS)
4,4'-Diisothiocyanostilbene-2,2'-Disulfonic Acid↗
Vasodilatation with pinacidil. Mode of action in rat resistance vessels.
Pinacidil is a newly developed antihypertensive vasodilator, proposed to belong to the new group of smooth muscle relaxants, the K+ channel openers. The in vitro effects of pinacidil on induced tone, smooth muscle membrane potential and 86Rb and 42K efflux from rat resistance vessels (internal diameter about 200 microns) were studied. Tone induced with noradrenaline was concentration-dependently inhibited by pinacidil. Responses to electrical field stimulation were also inhibited. However, tone induced with high K+ depolarisation, noradrenaline in the presence of high K+, caffeine-induced contractions and noradrenaline contractions in the presence of felodipine were little affected by pinacidil. Pinacidil caused concentration-dependent hyperpolarisation of the resting smooth muscle. Pinacidil caused only a small and transient increase of the 86Rb efflux rate constant, while the same concentrations of pinacidil produced a significant increase in the 42K efflux rate constant. Our results seem to indicate that the relaxant effect of pinacidil is the result of an increase in K+ permeability, thus causing hyperpolarisation and relaxation. The opened K+ channels appear to be selective for K+ over Rb+.
Animals↗
Calcium metabolism and structure in the peripheral vasculature: implications for hypertension.
This paper reviews in vitro investigations of resistance vessels taken from individuals with essential hypertension and from spontaneously hypertensive rats (SHRs). Unlike mesenteric resistance vessels from SHRs, the calcium sensitivity of subcutaneous resistance vessels from essential hypertensive patients is not increased, but decreased. However, like the resistance vessels from SHRs, the structure of the resistance vessels from the essential hypertensive patients (expressed in terms of their media thicknesses) was increased. In studies with SHRs, the increased media are found to be associated with cellular hyperplasia. If this also is true for the resistance vasculature of patients with essential hypertension, this may explain findings that antihypertensive treatment reduces blood pressure more readily than vascular structure. These findings could also be relevant as regards the rather limited effect of such treatment on hypertension-related diseases such as infarction.
Animals↗
Effect of changes in intracellular pH on the contractility of rat resistance vessels.
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Animals↗
Responses of femoral resistance vessels to angiotensin in vitro.
The effect of angiotensin II and angiotensin I on isolated rat resistance vessels (inner diameter ca. 200 micron) was investigated. Angiotensin II caused a contraction (ED50 = 0.58 +/- 0.17 X 10(-8) M) of rat femoral and cerebral arteries and to a lesser extent of mesenteric and renal arteries. However, all vessels showed strong tachyphylaxis on repeated stimulation with angiotensin II. Tachyphylaxis was avoided by inducing submaximal tone in the vessels with either K, noradrenaline or serotonin. The response to angiotensin II was inhibited by saralasin but not by captopril. Angiotensin I also caused contraction of the femoral arteries (ED50 = 2.68 +/- 0.32 X 10(-8) M). These responses were inhibited by captopril and saralasin. Functional removal of the endothelium had little effect on the contractile responses to either angiotensin I or II. These results indicate that there are functional receptors to angiotensin II in the resistance vessels of the rat and that, in the presence of tone (a more physiological condition), the vessels contract to angiotensin II without tachyphylaxis. In addition, angiotensin II may be formed from angiotensin I by the angiotensin converting enzyme which may be situated in the vessel wall as well as in the endothelium.
Angiotensin II↗
Contractile effects of tetradecapeptide renin substrate on rat femoral resistance vessels.
The effect of products of the renin-angiotensin system on the contractile response of rat resistance vessels (internal diameter approximately 200 microns) has been investigated. The vessels were isolated from the femoral bed and segments of the vessels were mounted on an isometric myograph. The vessels responded in a concentration-dependent manner to synthetic tetradecapeptide (TDP) renin substrate, angiotensin I (ANG I) and angiotensin II (ANG II), the responses to all these substances being inhibited by saralasin (0.1 mumol/l). The responses to ANG I, but not those to TDP renin substrate, were inhibited by captopril (1 mmol/l). In contrast, the non-specific protease inhibitor aprotinin had an inhibitory effect on responses to TDP renin substrate. The results suggest that TDP renin substrate is converted to ANG II by a process that does not involve metabolism of ANG I.
Angiotensin I↗
Endothelial-dependent relaxation in human subcutaneous resistance vessels.
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Acetylcholine↗