Increase In 42K Research Articles

Cromakalim- and RP 49356-induced 42K + and 86Rb + efflux in rat myometrium

Previous studies had failed to observe cromakalim-induced 42K + or 86Rb + efflux from the myometrium of the pregnant rat in contrast to positive findings in other smooth muscles. In the current study, in myometrium from the non-pregnant rat, cromakalim (10 μM) and RP 49356 ({(±)- N-methyl-2-(3-pyridyl)-tetrahydrothiopyran-2-(carbothioamide-1-oxide} 10 μM) induced small increases in 42K + or 86Rb + efflux but much less than did oxytocin (20 nM) or KCl (20 mM). The cromakalim-induced increase in 42K + efflux was enhanced 3.5-fold in the presence of KCl (20 mM) plus (+)- cis-diltiazem (3 μM), a property shared by RP 49356. Glibenclamide (10 μM) partially reduced the cromakalim-induced 42K + efflux, in the presence of KCl and (+)- cis-diltiazem, but did not affect the KCl-induced 42K + efflux. The data provides further support for the idea that cromakalim and RP 49356 are able to open potassium channels in rat myometrium. It would appear that their actions in this tissue are dependent on the extracellular K + concentration and/or membrane potential.

Cardiac ATP-sensitive K+ channels. Evidence for preferential regulation by glycolysis.

The ability of glycolysis, oxidative phosphorylation, the creatine kinase system, and exogenous ATP to suppress ATP-sensitive K+ channels and prevent cell shortening were compared in patch-clamped single guinea pig ventricular myocytes. In cell-attached patches on myocytes permeabilized at one end with saponin, ATP-sensitive K+ channels were activated by removing ATP from the bath, and could be closed equally well by exogenous ATP or substrates for endogenous ATP production by glycolysis (with the mitochondrial inhibitor FCCP present), mitochondrial oxidative phosphorylation, or the creatine kinase system. In the presence of an exogenous ATP-consuming system, however, glycolytic substrates (with FCCP present) were superior to substrates for either oxidative phosphorylation or the creatine kinase system at suppressing ATP-sensitive K+ channels. All three groups of substrates were equally effective at preventing cell shortening. In 6 of 38 excised inside-out membrane patches, ATP-sensitive K+ channels activated by removing ATP from the bath were suppressed by a complete set of substrates for the ATP-producing steps of glycolysis but not by individual glycolytic substrates, which is consistent with the presence of key glycolytic enzymes located near the channels in these patches. Under whole-cell voltage-clamp conditions, inclusion of 15 mM ATP in the patch electrode solution dialyzing the interior of the cell did not prevent activation of the ATP-sensitive K+ current under control conditions or during exposure to complete metabolic inhibition. In isolated arterially perfused rabbit interventricular septa, selective inhibition of glycolysis caused an immediate increase in 42K+ efflux rate, which was prevented by 100 microM glyburide, a known blocker of ATP-sensitive K+ channels. These observations suggest that key glycolytic enzymes are associated with cardiac. ATP-sensitive K+ channels and under conditions in which intracellular competition for ATP is high (e.g., in beating heart) that act as a preferential source of ATP for these channels.

Altered arterial ion transport and its reversal in aldosterone hypertensive rat.

Alterations in vascular ionic metabolism preceded the onset of elevated blood pressure in the aldosterone-treated rat. Increases in 42K+ and 36Cl- turnover in the aorta and femoral artery were detected as early as 1 wk after the start of aldosterone infusion. These vascular changes were completely reversed after a 3-wk recovery period. An increased sensitivity to the effect of norepinephrine (NE) on aortic 42K+ turnover was also observed prior to a significant rise in systolic blood pressure. Enhanced sodium transport was also found in vessels from hypertensive rats infused with aldosterone for 4 wk. The potassium-sensitive component of the 24Na+ efflux was significantly elevated in vessels from hypertensives. This increase in sodium pump activity appeared to compensate for an increase in passive membrane permeability to sodium in that cell sodium levels were not altered by aldosterone treatment. A more rapid increase in 42K+ turnover occurred in aorta from aldosterone-treated rats when extracellular calcium was removed. Therefore, an impaired ability of calcium-dependent processes to adequately stabilize the vascular smooth muscle membrane may contribute to altered vascular ion transport. It is concluded that altered vascular electrolyte metabolism is an important pathogenic mechanism in the development of aldosterone-induced hypertension in the rat.

Cation fluxes and volume regulation by human lymphocytes.

The ionic basis of volume regulation by human peripheral blood lymphocytes in hypotonic Tyrode's medium has been studied. The intracellular water space of lymphocytes increased to a maximum after 1 min in 0.68 X isotonic Tyrode's but returned to the isotonic value by 20 min at 37 degrees C. During this phase of volume regulation (1-20 min) both 42K+ efflux and 42K+ influx were stimulated severalfold, but the increase in 42K+ efflux exceeded the influx, resulting in a net loss of 20% of the lymphocyte K+. The increase in 42K+ efflux during the phase of cell shrinkage was unaffected by ouabain or by quinidine. Hypotonicity increased both the ouabain-sensitive (active) and ouabain-insensitive components of 42K+ influx by 76% and 123% respectively. Hypotonic shock stimulated 22Na+ influx by only 25%, but cell Na+ content was unchanged at 1 min and even decreased after 20 min. Thus active K+ influx and Na+ extrusion is increased by hypotonicity, but greater pumping cannot explain the net decrease in cell cations that leads to volume regulation. The 45Ca2+ uptake was not significantly changed by hypotonicity. Although volume regulation was abolished in a hypotonic high K medium, 42K+ efflux was still stimulated 2-fold by the reduction in tonicity. These findings support the hypothesis that volume regulation in hypotonic media occurs largely by a passive loss of cell K+, which results from a selective increase in membrane permeability to this ion. The increase in K+ permeability in hypotonic media is observed even in the absence of volume regulation by the cell.

Direct measurement of the increase in intracellular free calcium ion concentration in response to the action of complement.

1. The effect of rabbit anti-(pigeon erythrocyte) antibodies plus human complement on the concentration of intracellular free Ca2+ in sealed pigeon erythrocyte 'ghosts' was investigated with the photoprotein obelin. 2. The addition of human serum, as a source of complement, to 'ghosts' coated with antibody caused a rapid increase in intracellular free Ca2+ after a lag of 20-40 s, as detected by an increase in obelin luminescence. 3. The increase in obelin luminescence could not be explained by release of obelin into the medium. It was also Ca2+-dependent in that extracellular EGTA abolished the effect and intracellular EGTA inhibited it and required the complete terminal complex (C56789). No effect was seen with C5678. 4. The concentration of intracellular free Ca2+ before addition of complement was approx. 0.3 microM. This increased to a maximum of 5-30 microM after complement addition and then remained constant for at least 1-2 min. 5. Antibody plus complement induced a rapid increase in 42K+ efflux and an inhibition of cyclic AMP formation. 6. When partially purified complement components (C5b-9) were used in 'reactive lysis' it was possible to inhibit the release of macromolecules from pigeon erythrocyte 'ghosts' by extracellular EGTA. 7. It was concluded that the increase in intracellular free Ca2+ concentration caused by anti-cell antibody plus complement occurred before cell lysis and may be involved in the mechanism of complement-induced cell injury.

Effect of Concanavalin A on Intracellular K + and Na + Concentration and K + Transport of Human Lymphocytes

Incubation of human lymphocytes with ConA causes an increase in [Na +]; and a decrease in [K +];. This effect is not due to the experimental washing procedure, but is due to the ConAinduced increase in permeability which is not fully compensated by the increase in active transport. The ConA-induced increase in 42K + uptake consists of an increase in leak flux which is independent of [Na +] o, and of an increase in pump flux which is dependent on [Na +] 0. The increase in leak flux may be caused by increased membrane fluidity. The increase in pump flux may be produced by the increased [Na +]. and by a stimulation of Na +, K + ATPase.

Potasssium transport in human blood lymphocytes treated with phytohemagglutinin.

We have confirmed that phytohemagglutinin (PHA) rapidly enhances the uptake of potassium (K+) by human blood lymphocytes. PHA, however, did not produce an increase in lymphocyte K+ concentration. The apparent steady-state of cell K+ concentration despite the marked increase in uptake of 42K+ could be explained by either an increase in K+-K+ exchange or an increase in concentrative (active) K+ accumulation in association with an increase in the leak of K+ from the cell. We compared, therefore, the uptake of 42K+ with the decrement in cellular K+ content when active transport was inhibited by ouabain. These studies established that K+-K+ exchange was negligible in human blood lymphocytes and that the increase in 42K+ uptake after PHA treatment represented concentrative transport. Our studies did indicate that 42K+ exodus from PHA treated lymphocytes increased markedly from 19 to 38 mmol-1 cell water-1-h-1. Within the same time period K+ influx into PHA-treated lymphocytes increased from 20 to 38 mmol-1 cell water-1-h-1. Thus, PHA produces a marked increase in the permeability of the lymphocyte membrane to K+, and the increase in active K+ influx in PHA-treated lymphocytes may represent a homeostatic response by the membrane K+ transport system to the increase in K+ efflux. Increased K+ turnover was observed at the lowest concentrations of PHA which produced an observable increase in [3H]thymidine incorporation into DNA. Thus, PHA produces an increase in K+ permeability that closely parallels its mitogenic effect. The rapid increase in K+ influx preceding blastogenesis and mitogenesis is required, therefore, to maintain normal intracellular K+ concentration. An adequate intracellular K+ concentration is essential for the synthetic processes required for cell transformation or division.

EFFECTS OF MARINE TOXINS ON ELECTRICAL ACTIVITY AND K+ EFFLUX OF EXCITABLE MEMBRANES.

Two marine toxins clam poison and holothurin, have been tested on the Ranvier nodes of isolated myelinated nerve fiber and the monocellular electroplax preparation of Electrophorus electricus. Changes in action and resting potential were recorded with the air-gap technique or intracellular electrodes. Clam poison (10−8 M) blocks electrical activity without depolarization. Clam poison does not prevent the depolarization produced by acetylcholine or carbamylcholine. It has no effect on the increased 42K+ efflux produced by these compounds. Holothurin in a concentration of 10−5 M blocks electrical activity, and depolarizes the conducting membrane, an effect not prevented by tetracaine. Holothurin causes an initial increase in 42K+ efflux which steadily declines. The possible interactions of these toxins with the acetylcholine system are discussed.