Accumulation of Potassium and Calcium in Rat Myocardium Exposed to α‐1‐Adrenoceptor Stimulation

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The present study was undertaken to develop an improved and stabilized method for isolating cardiomyocytes from perfused rat heart. Different lots of the commercial collagenases used for isolating cardiomyocytes give variable results both with respect to the total cell yield and the percentage of elongated cells obtained. When trypsin was present both before and during collagenase treatment of the tissue, the performance of the collagenases was improved and stabilized, and a high and stable cell yield (7.5 x 10(6) cells per heart), and a high percentage of elongated cells (about 70%) was regularly obtained. The cells possessed alpha 1-adrenergic binding sites with binding properties (Bmax = 43.5 fmol/mg protein and Kd = 125.5 pmol/l) in agreement with values previously reported. The cells were able to respond functionally, as the cellular uptake of 86Rb+ increased by 18% after alpha 1-adrenoceptor stimulation with phenylephrine. These criteria indicate that the cells were well preserved during the isolation procedure.

Section: Discussionsupporting

confidence: 94%

Improved Isolation of Cardiomyocytes by Trypsination in Addition to Collagenase Treatment

Viko

Osnes

Sjetnan

et al. 1995

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“…All effects were prevented by the selective a-1adrenoceptor antagonist prazosin, thus confirming the involvement of this receptor type. An increase in intracellular potassium content based upon indirect measurements has previously been reported in heart (Ellingsen et al 1987;Hanem et al 1992). Direct measurements of intracellular potassium content and concentration in rat hearts after a-1-adrenoceptor stimulation have not to our knowledge been reported previously.…”

Section: Discussionmentioning

confidence: 69%

“…From electrophysiological studies on myocardial preparations, it has been established that outward potassium currents are reduced after a-I-adrenoceptor stimulation (Apkon & Nerbonne 1988;Wang et al 1991;Fedida et al 1993;Terzic et al 1993). Accumulation of potassium in hearts exposed to a-1 -adrenoceptor stimulation has been observed (Ellingsen et al 1987;Hanem et al 1992). In those two studies, however, the potassium accumulation was estimated indirectly by measuring the change in the arteriovenous differences.…”

mentioning

confidence: 99%

Increased Intracellular Potassium and Water Contents in Rat Heart after α‐1‐Adrenoceptor Stimulation

Hanem

Enger

Skomedal

et al. 1994

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Potassium accumulation in rat heart after alpha-1-adrenoceptor stimulation has previously been reported from indirect measurements. Here we present data on intracellular potassium content measured directly in the heart. Isolated rat hearts perfused in a non-recirculating system were exposed to alpha-1-adrenoceptor stimulation (5 x 10(-5) mol/l phenylephrine in the presence of 10(-6) mol/l timolol). 14C-Sucrose was used to estimate the extracellular space. From heart homogenates intracellular potassium, magnesium and cellular water contents were determined and the ion concentrations calculated accordingly. The intracellular magnesium content remained unchanged during all experimental conditions. alpha-1-Adrenoceptor stimulation evoked an increase in potassium content by 9% (4, 14; 95% confidence interval (CI), P = 0.0006). Due to an observed increase in intracellular water by 17% (9, 26; 95% CI, P = 0.0006), the potassium concentration apparently decreased by 8% (0.3, 15; 95% CI, P = 0.04). During partial inhibition of the Na+/K(+)-ATPase by 10(-5) mol/l ouabain, there was an increase in potassium content by 5% (1, 9; 95% CI, P = 0.008). There was, however, no significant increase in intracellular water in this situation. Calculated intracellular potassium concentration showed accordingly a slight increase. The effects upon potassium and water both in the absence and presence of ouabain were eliminated by the alpha-1-adrenoceptor blocker prazosin (10(-6) mol/l). alpha-1-Adrenoceptor stimulation apparently increased cellular dry weight by 10% (2, 18; 95% CI, P = 0.02). Changes in translocation of potassium and water must be considered as part of the alpha-1-adrenergic heart effects.

“…a,-Adrenoceptor stimulation decreases several outward K+-currents in rat ventricular cardiomyocytes (Fedida et al 1993;Terzic et al 1993). Studies using radioactive isotopes or measuring ionic concentrations, however, demonstrated both increased 86Rbf-influx (Hanem et al 1994a) and increased K+-influx (Ellingsen et al 1987a;Hanem et al 1992) and increased 86Rbf-efflux (Hanem et al 1993), resulting in net increased K+-uptake (Hanem et al 1994b). P-Adrenoceptor stimulation has also been shown to increase the K+-uptake in the heart (Ellingsen et al 1987a), and it was shown that this effect was mediated via stimulation of the P1-adrenoceptors (Ellingsen et al 1987b).…”

mentioning

confidence: 99%

α₁‐ and β‐Adrenoceptor‐Mediated Increase in ⁸⁶Rb⁺‐Uptake in Isolated Cardiomyocytes from Adult Rat Heart: Evidence for Interaction between the Two Receptor Systems

Viko

Osnes

Skomedal

1996

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The aim of the present investigation was to study the effect of alpha(1)- and beta-adrenoceptor stimulation, alone and in combination, on potassium uptake in isolated ventricular cardiomyocytes from adult rat heart, using the potassium analogue 86Rb+. The reliability of 86Rb+ as a potassium analogue was also investigated. Alpha(1)-, beta-, And combined adrenoceptor stimulation was achieved by using noradrenaline in the presence and absence of appropriate adrenoceptor antagonists. The uptake of 86Rb+ was found to increase linearly with time up to 20 min., both during basal and receptor-stimulated conditions. The basal uptake rate was about 0.18 ml/g protein x min. At 15 min. both alpha(1)-, beta- and combined adrenoceptor stimulation dose-dependently increased the 86Rb(+)-uptake with a -logEC50 of 7.05, 6.68 and 6.73, respectively. The maximal increase in these series achieved by 5 x 10(-5) mol/l noradrenaline was 29%, 24% and 41% above basal level, respectively. Comparison of the maximal effects in the same cell preparations, with the observed value for combined adrenoceptor stimulation in each experiment as 100%, gave a relative maximal increase in 86Rb(+)-uptake after separate alpha(1)-adrenoceptor stimulation of 67 +/- 8%, and of 68 +/- 6% after separate beta-adrenoceptor stimulation. The theoretically calculated value for combined adrenoceptor stimulation, if additivity, was 135 +/- 11%, which was significantly higher than the observed value (100%) (P = 0.026). The effect of noradrenaline was not limited by the maximal 86Rb(+)-uptake capacity, as 10(-5) mol/l forskolin increased the 86Rb(+)-uptake more than noradrenaline. Examining the reliability of 86Rb+ as potassium-analogue by combining 42K+ and 86Rb+ in the same experiments, showed that combined adrenoceptor stimulation dose-dependently increased both the 42K(+)- and 86Rb(+)-uptake with the same potency and to the same extent. Thus 86Rb+ is a reliable potassium-analogue for these effects. In conclusion both alpha(1)- and beta-adrenoceptor stimulation dose-dependently increased the cellular 86Rb(+)-uptake to the same extent and with the same potency. The observed maximal 86Rb(+)-uptake after combined adrenoceptor stimulation was significantly higher than the maximal effect after either form of separate receptor stimulation, but significantly lower than expected if the effects were purely additive. The results thus show inhibitory interaction between the two receptor systems.