1. The rate of heat production (heat rate) and isometric twitch tension of ventricular trabeculae isolated from guinea-pig heart were measured at 37°C in order to determine the relative contributions of actomyosin-ATPase, Ca2+-ATPase and Na+,K+-ATPase to myocardial energy metabolism. 2. The increase in heat rate recorded during isometric contractions at optimal length (contraction-related heat production) was 19-1 + 1P2 mW cm-3 at a stimulation rate of 2 Hz. The tension-time integral of individual contractions measured under the same conditions was 147 + 15 mN s cm2. 3. The heat production of the actomyosin-ATPase was determined by inhibiting the contractile proteins with 2,3-butanedione monoxime (BDM). Contraction-related heat production was reduced by 0-219 + 0 010 and the isometric tension-time integral was reduced by 0-288 + 0-016 in the presence of 1 mm BDM. From these data an estimate of 0-76 for the relative contribution of the actomyosin-ATPase to contraction-related heat production was derived.4. The heat production related to actomyosin-ATPase plus Ca2+-ATPase was studied by blocking Ca2+ influx into the myocardial cells with a solution containing 100 /M Ca2+ and 400 #M Ni2+. In this solution contraction-related heat production was reduced by 0'907 + 0-012. Comparison of this value with the component attributable to the actomyosin-ATPase yields an estimate of 0-15 for the relative contribution of the Ca2+-ATPase to contraction-related heat production. 5. The heat production related to the Na+,K+-ATPase in resting preparations was studied by blocking the sodium pump with 400 uM dihydro-ouabain (DHO). DHO produced a transient decrease in heat rate lasting 1-2 min, which was followed by a secondary increase. From the heat transient produced by DHO the heat rate related to the Na+,K+-ATPase in the steady state was extrapolated. The relative contribution of the sodium pump to resting heat production was estimated to be 0-17.6. The heat production related to the Na+,K+-ATPase in contracting preparations was studied by first blocking Ca2+ influx with 100 uM Ca2+ and 400,uM Ni2+, and then inhibiting the sodium pump with 400 uM dihydro-ouabain (DHO). The relative contribution of the sodium pump to contraction-related heat production extrapolated from these data was 0 10, which agreed well with the fraction of contraction-related heat production persisting after blockage of actomyosin-ATPase and Ca2+-ATPase (0 09).7. It is concluded that contraction-related heat production can be completely accounted for by the three major ATPases, and that under our experimental conditions actomyosinATPase, Ca2+-ATPase and Na+,K+-ATPase contribute about 76, 15 and 9%, respectively, to myocardial energy expenditure. 8. In cardiac muscle contracting under aerobic conditions the heat rate measured in the steady state is proportional to the rate of ATP hydrolysis, 1 1uW corresponding to about 11 pmol ATP s-5. From our heat and tension measurements the mechanical impulse per ATP molecule was calculated to be 0-027 pN s.

Monoquantal excitatory postsynaptic currents were recorded by means of a perfused macropatch electrode from 9 to 15 micro m stretches of crayfish neuromuscular junctions. The excitatory transmitter l-glutamate superfused to a terminal inhibits quantal release by activating autoreceptors [Parnas et al. (1996) Eur. J. Neurosci., 8, 116-126]. Substances related to glutamate that do not activate glutamatergic postsynaptic channels, but are substrates of glutamate transporters, elicited analogous inhibitions, e.g. l- and d-aspartate and some other glutamate transport blockers. As expected, all transport blockers prolonged synaptic currents. Blockers that bind to the transporter receptors but are not transported did not inhibit release, but prevented inhibition by the transport substrates. It appears that autoinhibition is elicited by transport of glutamate or its analogues. Transport into cells is powered by symport of three Na+. To block the transport step electrochemically, extracellular Na+ concentration was lowered to one-quarter, but this surprisingly left the inhibition of release by glutamate unaffected, showing inhibition to be associated to a step between binding and transport. After binding a substrate, glutamate transporters open a parallel Cl- channel. Replacement of extracellular Cl- prevented Cl- current, and release inhibition by glutamate or aspartate was blocked. It is suggested that the flow of Cl- across the cell membrane, after binding a transport substrate, mediates autoinhibition. We measured a related reduction of presynaptic action potentials.

To assess the effect of K adaptation on the electrolyte concentrations of renal tubular cells and on the concentration gradients across the luminal membrane, electron microprobe analysis was employed on freeze-dried cryosections of the renal cortex and on freeze-dried samples of tubular fluid in control and high-K rats. The measurements were performed in individual cells of the proximal and superficial distal tubule and on samples of tubular fluid obtained by free flow micropuncture from proximal and early and late distal collection sites. The ingestion of a potassium-rich diet for at least 10 days together with an acute potassium load of 0.4 mmol/kg/h led to a small increase in potassium concentration of about 7 mmol/kg wet weight (w.w.) in all cell types analysed. In distal convoluted tubule, connecting tubule and principal cells sodium concentration was markedly decreased by 4, 4, and 6 mmol/kg w.w., respectively, while no significant changes in sodium concentration were found in proximal tubule and intercalated cells. No consistent changes in cell chloride could be observed under K adaptation. Analysis of the tubular fluid samples showed that the K concentration gradient across the apical cell membrane of all distal tubular cell types investigated was diminished in the high-K rats. The concentration gradient for sodium entry, however, was clearly enhanced in the distal convoluted tubule, connecting tubule and principal cells.(ABSTRACT TRUNCATED AT 250 WORDS)

Studies were undertaken to define the effect of acute metabolic alkalosis (hypertonic sodium bicarbonate i.v.) on the chemical gradients for potassium, sodium and chloride across the apical membrane of individual renal tubule cells. Electron microprobe analysis was used on freeze-dried cryosections of the rat renal cortex to measure electrolyte concentrations in proximal tubule cells and in the various cell types of the superficial distal tubule. Analyses were also performed in fluid samples obtained by micropuncture from proximal and early and late distal collection sites. Compared with the appropriate controls (hypertonic sodium chloride i.v.), administration of sodium bicarbonate resulted only in small and mostly insignificant increases in cell potassium concentrations and induced only minor alterations in the cell/tubule fluid potassium concentration gradient for all cell types analysed. This observation suggests that under this condition factors other than an increase in cell potassium concentration are important in modulating potassium transfer across the apical membrane of potassium secreting cells. Nevertheless, since in alkalosis phosphorus and cell dry weight were decreased, and hence cell volume increased, in all but the intercalated cells, actually the potassium content of most tubular cells was higher under this condition. In comparison with animals infused with isotonic saline at low rates (hydropenic controls), infusion of either hypertonic sodium chloride or sodium bicarbonate led to a sharp increase in distal tubule fluid sodium concentrations and in the sodium concentrations of distal convoluted tubule, connecting tubule and principal cells, indicating that under both conditions the primary event causing enhanced transepithelial sodium absorption is stimulation of the sodium entry step.(ABSTRACT TRUNCATED AT 250 WORDS)

No abstract