F Karim

REGIONAL DIFFERENCES IN THE NEGATIVE INOTROPIC EFFECT OF ACETYLCHOLINE WITHIN THE CANINE VENTRICLE

1. Regional differences in the effects of ACh on sub‐epicardial, mid‐wall and sub‐endocardial cells of the dog left ventricle have been studied. 2. ACh produced a dose‐dependent, atropine‐sensitive negative inotropic effect that was greatest in sub‐epicardial cells and small or absent in sub‐endocardial cells. 3. In sub‐epicardial (but not sub‐endocardial) cells, ACh also resulted in a dose‐dependent decrease in action potential duration. The inotropic effect of ACh on sub‐epicardial cells was primarily the result of the decrease of action potential duration, because during trains of voltage clamp pulses the inotropic effect of ACh was reduced or abolished. At a holding potential of ‐80 mV, 10(‐5)M ACh decreased L‐type Ca2+ current by approximately 8% and this is thought to be responsible for the small inotropic effect during trains of pulses. 4. Although 4‐AP, a blocker of the transient outward current (I(to)), abolished the ‘spike and dome’ morphology of the sub‐epicardial action potential, it had little or no effect on the actions of ACh on sub‐epicardial cells. ACh had no effect on I(to) in sub‐epicardial cells in voltage clamp experiments. 5. ACh activated a Ba(2+)‐sensitive outward current (IK,ACh) in sub‐epicardial cells, but little or no such current in sub‐endocardial cells. In sub‐epicardial cells, ACh also inhibited the inward rectifier current, IK,1. 6. It is concluded that in left ventricular sub‐epicardial cells, ACh activates IK,ACh. This results in a shortening of the action potential and, therefore, a negative inotropic effect. In subendocardial cells, ACh activates little or no IK,ACh and, therefore, it has little or no negative inotropic effect. This may result from a regional variation in the expression of the muscarinic K+ channel.

Appearance of adenosine in venous blood from the contracting gracilis muscle and its role in vasodilatation in the dog.

1. In dogs anaesthetized with sodium pentobarbitone and artificially ventilated, the gracilis muscles were vascularly isolated and perfused at a constant flow rate of 51.2 +/‐ 9.8 ml min‐1 100 g‐1 muscle tissue (183 +/‐ 17.8% of resting blood flow; mean +/‐ S.E.; n = 13). 2. Electrical stimulation of the cut peripheral end of the obturator nerve (6 V, 4 Hz) resulted in muscle contraction (658 +/‐ 118 g 100 g‐1 force after 5 min), and an immediate decrease in arterial perfusion pressure from 179 +/‐ 15.7 mmHg to 87 +/‐ 10.0 mmHg (51.4 +/‐ 4.5% decrease in vascular resistance after 2 min of contraction). Venous oxygen tension decreased from 69.2 +/‐ 5.1 mmHg to 18.5 +/‐ 1.4 mmHg (n = 6). These values did not significantly alter during the remaining period of stimulation (10‐20 min). 3. The concentration of adenosine in arterial plasma did not change significantly during muscle contraction (137 +/‐ 23 nM; n = 10). However, the adenosine concentrations in venous plasma showed a significant (P less than 0.01) increase from a control value of 164 +/‐ 55 nM to 455 +/‐ 77 nM (n = 9) after 5 min of muscle contraction and remained high during the rest of the 20 min contraction. In six of the dogs adenosine concentrations were determined after 1 and 3 min of contraction and showed a smaller but statistically significant (P less than 0.05) rise in venous concentration. 4. During infusion of adenosine into the artery to give plasma concentrations between 0.3 microM and 1 mM, 72.6 +/‐ 2.1% (n = 29) of the infused adenosine was taken up by the tissues before it reached the vein. Comparison of vasodilatation and venous adenosine concentrations during adenosine infusion and muscle contractions showed that the released adenosine could contribute about 15% to the total vasodilatation after 1 min and about 40% between 5 and 20 min of contractions. Released adenosine could contribute about 80% to the vasodilatation that remained 5 min after the withdrawal of stimulation. Arterial perfusion pressure took 22 min to return to control, whereas adenosine release had fallen to zero within 10 min. 5. These data suggest that the released adenosine could contribute to exercise hyperaemia, but is unlikely to be the main factor, particularly in the initial stage.

Attenuation of exercise vasodilatation by adenosine deaminase in anaesthetized dogs

1. In dogs anaesthetized with sodium pentobarbitone and artificially ventilated, the gracilis muscles were vascularly isolated and perfused at a constant flow of 28.4 +/‐ 4.6 ml min‐1 (100 g muscle tissue)‐1 (99.8 +/‐ 4.5% of maximum free flow, means +/‐ standard error of the mean (S.E.M.), n = 9). 2. Three to five minutes of electrical stimulation of the cut peripheral end of the obturator nerve (4 Hz, 6 V, 0.2 ms) resulted in muscle contraction (0.61 +/‐ 0.14 kg (100 g)‐1 during solvent infusion and 0.56 +/‐ 0.10 kg (100 g)‐1 during intra‐arterial adenosine deaminase infusion (50 U min‐1) and an immediate decrease in arterial perfusion pressure from 184.5 +/‐ 8.1 mmHg to 148.2 +/‐ 5.7 mmHg (18.7 +/‐ 3.4% decrease) during solvent infusion, and from 193.5 +/‐ 7.16 to 142.0 +/‐ 10.2 mmHg (25.4 +/‐ 6.1% decrease) during adenosine deaminase infusion 10 s after the commencement of muscle stimulation. After about 5 min of muscle contractions, the arterial perfusion pressure decreased to 120.8 +/‐ 7.8 mmHg (32.9 +/‐ 5.8% decrease) during solvent infusion, and to 152.8 +/‐ 11.2 mmHg (20.9 +/‐ 5.3% decrease) during adenosine deaminase infusion (i.e. 37.9 +/‐ 6.2% attenuation of the fall in arterial perfusion pressure). The time taken for 90% recovery of the arterial perfusion pressure was 72.1 +/‐ 10.9 s during solvent infusion, and 51.5 +/‐ 9.3 s during adenosine deaminase infusion (P less than 0.05). 3. Adenosine (2 x 10(‐3) mol l‐1) infusion in the resting muscle during solvent infusion (final concentration in arterial blood 1.3 x 10(‐4) +/‐ 6.0 x 10(‐5) mol l‐1) resulted in a 34.8 +/‐ 7.2% fall in arterial perfusion pressure but a fall of only 7.2 +/‐ 1.8% during adenosine deaminase infusion (50 U min‐1; P less than 0.05; n = 5) indicating that adenosine deaminase infused at 50 U min‐1 was more than adequate to metabolize endogenous adenosine produced during muscle contractions. 4. These data suggest that adenosine contributes about 40% to the sustained‐exercise vasodilatation under constant high‐flow conditions and also in post‐exercise vasodilatation, but does not contribute to the initiation of exercise vasodilatation.

The effects of stimulating carotid chemoreceptors on renal haemodynamics and function in dogs.

1. Dogs were anaesthetized with chloralose and artificially ventilated. The carotid chemoreceptors were stimulated by changing the perfusion of vascularly isolated carotid sinus regions from arterial to venous blood. The mean carotid sinus pressure and the mean arterial blood pressure were held constant at 124 +/‐ 3 and 122 +/‐ 3 mmHg, respectively. Both vagosympathetic trunks were sectioned in the neck and propranolol (17 micrograms kg‐1 min‐1 I.V.) and gallamine triethiodide (0.2‐2.0 mg kg‐1 30 min‐1 I.V.) were infused. Renal blood flow was measured by an electromagnetic flow probe, glomerular filtration rate by creatinine clearance, sodium excretion by flame photometry and solute excretion by osmometry. 2. In sixteen tests in thirteen dogs perfusion of the carotid sinus regions with venous blood resulted in significant decreases in renal blood flow from 271 +/‐ 24 to 198 +/‐ 21 ml min‐1 100 g‐1 renal mass; glomerular filtration rate from 41.0 +/‐ 4.8 to 22.1 +/‐ 3.1 ml min‐1 100 g‐1; filtration fraction from 0.25 +/‐ 0.02 to 0.19 +/‐ 0.02; urine flow from 0.48 +/‐ 1.0 to 0.21 +/‐ 0.03 ml min‐1 100 g‐1; sodium excretion from 18.1 +/‐ 4.1 to 12.9 +/‐ 4.2 mumol min‐1 100 g‐1; and osmolar excretion 327 +/‐ 42 to 171 +/‐ 26 mu osmol min‐1 100 g‐1. The right atrial pressure did not change significantly from 4.6 +/‐ 1.2 cmH2O. 3. In seven dogs, tying renal sympathetic nerves abolished all the responses except that of sodium excretion which was now reversed; sodium excretion increased from 68 +/‐ 19 to 116 +/‐ 38 mumol min‐1 100 g‐1 without significant change in right atrial pressure from 7.4 +/‐ 1.9 cmH2O. Crushing the carotid bodies, however, abolished all the responses. 4. The results show that carotid chemoreceptor stimulation can cause significant reflex effects on renal haemodynamics and function which are mediated via renal sympathetic nerves. They also show that the chemoreceptor stimulation can cause natriuresis in the absence of haemodynamic changes, in the denervated kidney, presumably via a humoral factor.

Modification of carotid chemoreceptor-induced changes in renal haemodynamics and function by carotid baroreflex in dogs.

1. Mongrel dogs were anaesthetized with thiopental sodium and chloralose and artificially ventilated. The carotid sinus regions were vascularly isolated and perfused either with arterial or mixed (arterial and venous) blood (PO2, 44.2 +/‐ 3.3 mmHg, mean +/‐ S.E.M.) to stimulate the chemoreceptors. Cervical vagosympathetic trunks were ligated and atenolol (2 mg kg‐1, I.V.) was given in all dogs and gallamine triethiodide (3 mg kg‐1 h‐1, I.V.) was given in two dogs. Renal blood flow was measured by an electromagnetic flowmeter, glomerular filtration rate by creatinine clearance, sodium excretion by flame photometry and solute excretion by osmometry. The viability of the preparations was tested by recording total vascular capacitance responses to stimulation of carotid baro‐ and chemoreceptors. 2. In eleven tests in seven dogs at a constant aortic pressure of 88.9 +/‐ 2.6 mmHg stimulation of carotid chemoreceptors at a high carotid sinus pressure (194.0 +/‐ 3.6 mmHg) resulted in significant increases in urine flow of 22.8 +/‐ 3.0%, urinary sodium excretion of 30.7 +/‐ 5.2%, fractional sodium excretion of 35.3 +/‐ 18.6%, osmolar excretion of 17.5 +/‐ 4.1% and a decrease in free water clearance of 30.8 +/‐ 3.1% without significant changes in urinary sodium concentration, renal blood flow, glomerular filtration rate, and filtration fraction. 3. In seventeen tests in these seven dogs at a constant aortic pressure of 94.0 +/‐ 2.2 mmHg, stimulation of carotid chemoreceptor at a low carotid sinus pressure (72.0 +/‐ 1.3 mmHg) resulted in significant decreases in renal blood flow of 10.6 +/‐ 2.5%, glomerular filtration rate of 19.6 +/‐ 6.8%, filtration fraction of 13.2 +/‐ 5.5%, urine flow of 23.4 +/‐ 4.1%, urinary sodium concentration of 20.3 +/‐ 4.1%, urinary sodium excretion of 38.5 +/‐ 4.6%, fractional sodium excretion of 20.2 +/‐ 7.7%, osmolar excretion of 23.9 +/‐ 4.0% and an increase in free water clearance of 23.1 +/‐ 2.5%. 4. The results show that moderate stimulation of carotid chemoreceptors at a low carotid sinus pressure, when the activity in renal nerves is high and blood volume is low, can produce significant reflex decreases in renal haemodynamic and functional variables. However, at a high carotid sinus pressure when the renal sympathetic activity is low and blood volume is high, carotid chemoreceptor stimulation produces diuresis and natriuresis but no change in renal haemodynamics.

Sympathetic nerves in the mediation of renal response to localized stimulation of atrial receptors in anaesthetized dogs.

1. Dogs were anaesthetized with chloralose and artificially ventilated. Localized stimulation of left atrial receptors for 23‐25 min was achieved by distension of three small balloons at the pulmonary vein‐atrial junctions and one in atrial appendage. Renal blood flows were measured by electromagnetic flow probes, glomerular filtration rate by creatinine clearance, urinary sodium excretion by flame photometry and solute excretion by osmometry. The mean aortic pressure was held constant at 92.2 +/‐ 2.4 mmHg (mean +/‐ S.E.M., n = 27) by means of a pressure bottle connected to the aorta and beta‐adrenergic receptor activity was blocked by continuous infusion of propranolol (17 micrograms kg‐1 min‐1, I.V.). 2. In twelve dogs stimulation of left atrial receptors resulted in significant increases of 11.8 +/‐ 2.4% (P less than 0.001) in renal blood flow; 32.5 +/‐ 7.2% (P less than 0.001) in glomerular filtration rate; 19.5 +/‐ 5.0% (P less than 0.005) in filtration fraction: 36.3 +/‐ 9.0% (P less than 0.001) in urine flow: 32.7 +/‐ 9.2% (P less than 0.005) in sodium excretion: 36.6 +/‐ 9.9% (P less than 0.005) in osmolar excretion and a decrease of 31.3 +/‐ 11.2% (P less than 0.025) in free water clearance. Left atrial pressure and heart rate did not change significantly. In eight of the dogs ligation of the renal nerves resulted in similar changes in all of the renal variables; subsequent stimulation of atrial receptors did not cause significant changes in the renal variables. 3. In five additional dogs, in which heart rate and aortic pressure were allowed to change, stimulation of left atrial receptors for the same period resulted in significant increases in heart rate (4.3 +/‐ 0.7%. P less than 0.001) and mean aortic pressure (2.0 +/‐ 0.6%, P less than 0.025). Under this condition both the intact right kidneys and the denervated left kidneys showed significant responses in urine flow, sodium excretion, osmolar excretion and free water clearance. 4. The results show that the renal sympathetic nerves mediate the primary renal responses to atrial receptor stimulation, at least in the short term. The influence of any humoral factor in this reflex seems to be secondary to changes in heart rate and systemic blood pressure, possibly via arterial baroreceptors.

The role of adenosine in functional hyperaemia in the coronary circulation of anaesthetized dogs.

1. The aim of this investigation was to determine the contribution of adenosine to coronary active hyperaemia in the dog denervated heart by using adenosine deaminase. 2. Beagles were anaesthetized with thiopentone sodium (500 mg, I.V.) and chloralose (100 mg kg‐1, LV.) and artificially ventilated. The hearts were denervate by bilateral cervical vagotomy and cardiac sympathectomy. Blood samples were collected from the coronary sinus via a cannula passed through the right external jugular vein. The anterior descending or circumflex branch of the left coronary artery was cannulated and perfused with blood from the left subclavian artery under systemic blood pressure through an electromagnetic flow probe and a perfusion circuit. The heart was paced (3 V, 0.2 ms and a suitable frequency) via two electrodes attached to the right atrium from 109 +/‐ 7.3 to 170 +/‐ 9.8 beats min‐4 (means +/‐ S.E.M.) for 3‐4 min, first during an infusion of the solvent, and then during an infusion of a solution of adenosine deaminase (5 U kg‐1 min‐1) into the circuit. 3. In seventeen tests in eight dogs, infusion of adenosine deaminase did not cause a significant change in the basal coronary blood flow nor in the immediate increase (within 10s) in blood flow induced by pacing the heart from its basal rate to 170 beats min‐1. However, adenosine deaminase did cause a significant attenuation by 58 +/‐ 5.2% (P < 0.05) of the increase in coronary blood flow induced at 3‐4 min of pacing from 31 +/‐ 4.6 to 43 +/‐ 5.8 ml min‐1 (100 g cardiac tissue)‐1. Concomitantly, the pacing‐induced increase in coronary vascular conductance (from 0.41 +/‐ 0.08 to 0.54 +/‐ 0.12 ml min‐1 (100 g)‐1 mmHg‐1) was reduced by 75 +/‐ 6.6% (P < 0.02) and the increase in myocardial O2 consumption (from 13 +/‐ 3.5 to 21 +/‐ 4.2 ml min‐1 (100 g)‐1) was reduced by 50 +/‐ 12% (P < 0.05) but without significant changes in oxygen extraction or myocardial contractility. 4. The results show that although adenosine is unlikely to play a significant role in the regulation of the basal coronary blood flow, it can play a major role in the coronary active (functional) hyperaemia induced by atrial pacing to a high rate in the denervated heart of anaesthetized dogs.

The reflex effects of changes in carotid sinus pressure upon renal function in dogs.

In chloralose‐anaesthetized and artificially ventilated dogs, the carotid sinuses were vascularly isolated and perfused with arterial blood. Mean aortic pressure was held constant at 100 +/‐ 2 mmHg (mean +/‐ S.E. of mean, n = 19) by means of a pressure bottle connected to the aorta. Both vagus nerves were sectioned in the neck and propranolol hydrochloride (0.5 mg kg‐1) was administered every 30 min. The left renal blood flow was measured by an electromagnetic flowmeter (wrap‐round probe), glomerular filtration rate by creatinine clearance and urinary sodium by flame photometry. Decreasing pressure in the isolated carotid sinuses from 186 +/‐ 10 to 63 +/‐ 5 mmHg resulted in significant decreases in renal blood flow from 281 +/‐ 35 to 177 +/‐ 30 ml min‐1 100 g‐1 renal mass; glomerular filtration rate from 40.0 +/‐ 7.8 to 12.3 +/‐ 4.4 ml min‐1 100 g‐1; urine flow from 0.31 +/‐ 0.05 to 0.12 +/‐ 0.03 ml min‐1 100 g‐1 and sodium excretion from 21.7 +/‐ 7.2 to 8.2 +/‐ 3.0 mumol min‐1 100 g‐1. Increasing carotid sinus pressure back to 188 +/‐ 11 mmHg resulted in increases in all the variables to values not significantly different from their initial values. Tying renal sympathetic nerves at low carotid sinus pressure (73 +/‐ 11 mmHg) caused an increase in all of the variables. After denervation there was no response to changes in carotid sinus pressure. These results show that changes in carotid sinus pressure can result in significant reflex effects on renal function and that these effects are mediated by renal sympathetic nerves.

Effects of adenosine and its analogues on the perfused hind limb artery and vein of anaesthetized dogs

1. The effects of infusion of adenosine and its analogues on arterial and venous resistance have been studied in the vascularly and sympathetically isolated hind limb of chloralose‐anaesthetized dogs. Resistance changes have been assessed by monitoring changes in perfusion pressures at constant flow through the femoral artery and metatarsal vein.

Primary effects of carotid chemoreceptor stimulation on gracilis muscle and renal blood flow and renal function in dogs.

1. In chloralose‐anaesthetized and artificially ventilated dogs, the carotid sinus regions were vascularly isolated and perfused either with arterial or mixed (arterial and venous) blood (partial pressure of O2 (PO2) 43.8 +/‐ 2.4 mmHg, mean +/‐ S.E.M. n = 14) to stimulate the carotid chemoreceptors. The carotid sinus pressure was held constant at 142.0 +/‐ 2.8 mmHg. Measurements were made of renal and gracilis muscle blood flow by wrap‐round electromagnetic flow probes placed around the renal and gracilis arteries, glomerular filtration rate by creatine clearance, urinary sodium excretion by flame photometry and solute excretion by osmometry. 2. In ten dogs, with intact cervical vagosympathetic trunks, carotid chemoreceptor stimulation produced significant increases in aortic pressure (AoP) of 12.7 +/‐ 1.1% (n = 10, P < 0.001), in glomerular filtration rate (GFR) of 14.7 +/‐ 4.1% (P < 0.001), urine flow rate (V) of 16.5 +/‐ 3.5% (P < 0.002), in urinary sodium excretion (UNaV) of 17.5 +/‐ 2.5% (P < 0.005) and in urinary osmolar excretion (UosmV) of 13.2 +/‐ 2.2% (P < 0.001), but a significant decrease in renal blood flow (RBF) of 5.8 +/‐ 1.8% (P < 0.02). In six of these dogs in which gracilis muscle blood flow (MBF) was also recorded, carotid chemoreceptor stimulation caused significant increases in AoP of 12.8 +/‐ 1.4% (n = 6, P < 0.001) and in MBF of 10.0 +/‐ 1.6% (P < 0.002), and a small but significant decrease in RBF of 3.6 +/‐ 1.5% (P < 0.02). 3. In fourteen dogs, with sectioned cervical vagosympathetic trunks, carotid chemoreceptor stimulation produced increases in AoP of 22.0 +/‐ 2.6% (n = 14, P < 0.001), in GFR of 36.9 +/‐ 4.2% (P < 0.001), in V of 30.1 +/‐ 4.4% (P < 0.001), in UNaV of 31.4 +/‐ 5.3% (P < 0.001), and in UosmV of 25.7 +/‐ 5.8% (P < 0.001). However, it produced a greater decrease in RBF of 10.5 +/‐ 1.9% (P < 0.001). In ten of these dogs, where MBF was recorded, carotid chemoreceptor stimulation caused greater increase in AoP of 22.4 +/‐ 3.0% (n = 10, P < 0.001) and in MBF of 32.8 +/‐ 3.7% (P < 0.001), and a greater decrease in RBF of 9.8 +/‐ 1.9% (P < 0.001).(ABSTRACT TRUNCATED AT 400 WORDS)