Blockade of the Na+-Ca2+ exchanger is more efficient than blockade of the Na+-H+ exchanger for protection of the myocardium from lethal reperfusion injury

Since the Na⁺-H⁺ exchanger (NHE) is not the only pathway of Na⁺ influx into cardiomyocytes during ischemia/reperfusion, we hypothesized that blockade of Na⁺-Ca²⁺ exchanger (NCX) may be a more efficient strategy than is NHE inhibition for protecting the myocardium from infarction. To test this hypothesis, we compared KB-R7943 (KBR), a novel selective NCX blocker, with cariporide, a selective NHE blocker, with regard to their protective effects against infarction. In isolated rabbit hearts, infarction was induced by 30-min global ischemia/2-h reperfusion, and infarct size was determined by tetrazolium staining and expressed as a percentage of area at risk (%IS/AR). Hearts received no drugs, or were infused with cariporide (1 M) for 10 min or KBR (0.3 or 10 M) for 5 min before ischemia or after the onset of reperfusion. Protein level of NCX was assessed by Western blotting. Cariporide infusion before ischemia significantly reduced %IS/AR from 63.9 ± 2.9% to 20.2 ± 3.0%, but its infusion upon reperfusion failed to achieve a significant protection (%IS/AR = 53.8 ± 4.6%). In contrast, KBR infusion similarly reduced infarct size both when infused before ischemia (%IS/AR = 33.3 ± 6.3% and 21.9 ± 4.7% by 0.3 and 10 M KBR, respectively) and when infused for only 5 min after reperfusion (%IS/AR = 35.3 ± 7.1% and 31.5 ± 2.1% by 0.3 and 10 M KBR, respectively). Protein levels of NCX after 30-min ischemia and 30-min ischemia/30-min reperfusion were similar to baseline values in both untreated controls and hearts treated with 0.3 M KBR upon reperfusion. These results suggest that lethal reperfusion injury is more efficiently suppressed by blockade of the NCX than by blockade of the NHE.     

DA1 Receptor Mediated Regulation of Na+-H+ Antiport Activity in Rat Renal Cortical Brush Border Membrane Vesicles

Our previous studies indicate that dopamine (DA) plays an important role in regulating renal sodium (Na⁺) metabolism during high Na⁺ intake, and that DA₁ receptors are involved in natriuretic response to acute volume expansion. It has also been shown that in addition to the changes in renal hemodynamics, the natriuretic response produced by exogenously administered DA and DA₁ receptor agonists appears to be due to alterations in renal tubular sodium transport mechanisms. This study was designed to investigate the DA₁ receptor-mediated changes in Na⁺-H⁺ antiport activity in tubular brush border membranes of rat kidney. The Na⁺-H⁺ antiport activity, measured as the amiloride-sensitive Na⁺ influx in BBMV, was inhibited by 37%, 46%, 33%, and 42% by 1 μM DA, SKF 82958, SKF 38393, and fenoldopam respectively. The DA₁ antagonist SCH 23390 increased the antiport activity when given alone, while when administered with an agonist it attenuated the effects of the agonist on the antiporter. DA₂ agonists and antagonists failed to affect the antiport activity. These results indicate that the inhibitory effects of DA and DA receptor agonists on Na⁺-H⁺ antiport activity in renal cortical BBMV were mediated by the DA₁ receptors.     

Role of Na+-H+-antiport in restitution of isolated guinea pig gastric epithelium after superficial injury

In addition to its pH_i regulatory function Na⁺-H⁺-antiport is also involved in volume regulation of epithelial cells, particularly in neutral conditions. It is also known that the antiport is activated after ligand binding following growth factor receptor activation. The aim of the present study was to evaluate the role of the antiport in restitution of gastric mucosa and whether its activity is dependent on the type of superficial injury. Therefore the fundic epithelium of guinea pig stomach was perfused in an Ussing chamber in neutral conditions. Na⁺-H⁺- and Cl^–-HCO ₃ ^– -antiports were inhibited with 1.0 mM amiloride, 1.0 mM SITS, or with HCO ₃ ^– removal and Na⁺-K⁺-2Cl^2–-cotransporter with 0.3 M furosemide during 4 hr of restitution after superficial injury induced either by 1.25 M NaCl or by 1.0% Triton. Luminal exposure of the epithelium to amiloride had no effect on restitution but serosal application abolished the process completely. The inhibitory effect of amiloride was similar after both NaCl and Triton injury. The inhibition of Cl^–-HCO ₃ ^– -antiport with SITS interfered with the process as well, while HCO ₃ ^– removal had no significant inhibitory effect, nor did the inhibition of Na⁺-K⁺-2Cl^–-cotransporter. The morphologic findings were in accordance with the electrophysiologic measurements in each pair of tissues. It is concluded that the Na⁺-H⁺-antiport is essential for the epithelial cells during restitution even in neutral conditions, but a functional Cl^–-HCO ₃ ^– -antiport is also required. The activity of Na⁺-H⁺-antiport is sensitive to basolateral amiloride and is necessary regardless of the type of chemical injury.This study was supported by grants from Astra Co., the Orion Corporation Research Foundation, the Sigrid Juselius Foundation, the Academy of Finland, the Research Foundation of Gastrointestinal Diseases, the Clinical Research Institute of Helsinki University Central Hospital, Helsinki, Finland.     

Effects of Na+-H+ exchange blocker amiloride on left ventricular remodeling after anterior myocardial infarction in rats

Summary We investigated the effects of amiloride, a Na⁺-H⁺ exchange blocker, on ventricular remodeling in an infarcted rat model. In the amiloride group, the left descending coronary artery was ligated and rats were given amiloride (1 mg/kg/day, n=11) in their drinking water for 4 weeks. In the control group, rats were given water for 4 weeks (n=8) after myocardial infarction. The rats were killed on day 28. Both the ratio of heart weight to body weight and that of left ventricular weight to body weight were significantly less in the amiloride group (p<0.05). The diameter of a myocardial fiber in the region adjacent to the operated area was significantly reduced in the amiloride group compared with the control group (p<0.05). Left ventricular cavity dimension was significantly smaller in the amiloride group than that in control group (p<0.05). Our findings suggest that amiloride prevents ventricular remodeling after myocardial infarction.     

Gap junction and Na+-H+ exchanger alternations in fibrillating and failing atrium

Atrial fibrillation (AF) frequently complicates congestive heart failure (CHF). The current study examines whether patients with AF and CHF have alternation in atrial connexins (Cxs) and Na⁺–H⁺ exchanger isoform 1 (NHE1). Left atrium was obtained from 29 patients of patients undergoing mitral valve replacement. The connexin 40 (Cx40) protein was significantly higher in AF and sever HF, whereas connexin 43 (Cx43) protein were markedly increased in AF and mild HF but downregulated in the patients with AF and sever HF. NHE activity is significantly greater in AF and sever HF with highest level of NHE1 protein. These change may induce a direct effect on the progressive deterioration of cardiac function in the fibrillation and failure heart.     

ATP-dependent Ca2+ Transport and Na+/Ca2+ Exchange System in Renal Basolateral Plasmamembranes of Spontaneously Hypertensive Rats

A common abnormality of cellular Ca²⁺ handling in most tissues of spontaneously hypertensive rats (SHR) has been suggested. Therefore we investigated the ATP-dependent Ca²⁺ transport and Na⁺/Ca²⁺ exchange system in basolateral membrane vesicles (BLMV) of renal cortices from SHR and normotensive Wistar-Kyoto rats (WKY) at 12 and 20 weeks of age. In WKY the maximal transport rate of the ATP-dependent Ca²⁺ transport was 5.7 nmol/min/mg prot with an affinity for Ca²⁺ of 0.14 µM. These values were not significantly different in SHR at both ages studied. High concentrations of Na⁺ inhibited ATP-dependent Ca²⁺ uptake by 40% in BLMV of SHR and WKY. Low concentrations of Na⁺ stimulated ATP-dependent Ca²⁺ transport 20% in both rats. These findings suggest equal Na⁺/Ca²⁺ exchange activity in WKY and SHR. The present study failed to show a significant change in ATP-dependent Ca²⁺ transport and Na⁺/Ca²⁺ exchange activity in renal BLMV in SHR, suggesting that the Ca²⁺ homeostasis of the cortical cells is normal in SHR as far as the plasmamembrane is concerned.     

Endothelin-1 induced hypertrophic effect in neonatal rat cardiomyocytes: involvement of Na+/H+ and Na+/Ca2+ exchangers

Endothelin-1 (ET-1) is a potent agonist of cell growth that also stimulates Na(+)/H(+) exchanger isoform 1 (NHE-1) activity. It was hypothesized that the increase in intracellular Na(+) ([Na(+)](i)) mediated by NHE-1 activity may induce the reverse mode of Na(+)/Ca(2+) exchanger (NCX(rev)) increasing intracellular Ca(2+) ([Ca(2+)](i)) which in turn will induce hypertrophy. The objective of this work was to test whether the inhibition of NHE-1 or NCX(rev) prevents ET-1 induced hypertrophy in neonatal rat cardiomyocytes (NRVMs). NRVMs were cultured (24 h) in the absence (control) and presence of 5 nmol/L ET-1 alone, or combined with 1 mumol/L HOE 642 or 5 mumol/L KB-R7943. Cell surface area, (3)H-phenylalanine incorporation and atrial natriuretic factor (ANF) mRNA expression were increased to 131 +/- 3, 220 +/- 12 and 190 +/- 25% of control, respectively (P < 0.05) by ET-1. [Na(+)](i) and total [Ca(2+)](i) were higher (8.1 +/- 1.2 mmol/L and 636 +/- 117 nmol/L, respectively) in ET-1-treated than in control NRVMs (4.2 +/- 1.3 and 346 +/- 85, respectively, P < 0.05), effects that were cancelled by NHE-1 inhibition with HOE 642. The rise in [Ca(2+)](i) induced by extracellular Na(+) removal (NCX(rev)) was higher in ET-1-treated than in control NRVMs and the effect was prevented by co-treatment with HOE 642 or KB-R7943 (NCX(rev) inhibitor). The ET-1-induced increase in cell area, ANF mRNA expression and (3)H-phenylalanine incorporation in ET-1-treated NRVM were decreased by NHE-1 or NCX(rev) inhibition. Our results provide the first evidence that NCX(rev) is, secondarily to NHE-1 activation, involved in ET-1-induced hypertrophy in NRVMs.     

Decreased Na+ K +ATPase Activity in the Aortic Endothelium and Smooth Muscle of the Spontaneously Hypertensive Rats

The activity of Na⁺ K⁺ ATPase in the endothelium and smooth muscle of the aortae of normotensive and hypertensive rats was investigated. The enzyme activity in the endothelium and smooth muscle of the spontaneously hypertensive rats (SHR) was 2.15 ± 0.48 and 12.98±0.99 respectively. These values were significantly lower (P<0.05) than the enzyme activity in the corresponding tissues (10.10 ± 1.78 for endothelium, 20.77 ± 2.54 for smooth muscle) of the normotensive Wistar Kyoto (WKY) rats. However, with the low blood pressure spontaneously hypertensive rats (LBP-SHR) i.e. in those animals whose blood pressures were below 150 mm Hg, the enzyme activity in both tissues was not significantly different from those of the WKY. Since Na⁺ K⁺ ATPase is coupled to the sodium-potassium pump whose activity affects the functions of other pumps, the results indicate that the development of high blood pressure in the SHR may be related to an alteration in the transport of cations across the cell membrane.     

Evidence for apical Na+/H+ exchanger in bovine main pancreatic duct

The finding of a high Pco ₂ in basally secreted pancreatic juice of man and dog raises the hypothesis of proton secretion from ductal epithelial cells presumably through a Na⁺/H⁺ exchanger. To test this possibility, H⁺ luminal secretion and Na⁺ movements were measuredin vitro on samples of bovine pancreatic ducts mounted in Ussing-type chambers. The rate of luminal acidification measured by the pH stat method, using bicarbonate-free media gassed with 100% O₂, reached 2.75 Eq/cm²/hr. Proton secretion was blocked in the presence of 1 mM amiloride or in the absence of Na⁺ (replaced by choline) in the mucosal solution. Study of transepithelial²²Na fluxes in short-circuited tissue, bathed on both sides by control Ringer solution, gassed by 95% O₂-5% CO₂ demonstrated a net sodium transport from the mucosal to the interstitial side of the duct (net²²Na flux=3.23±0.8 Eq/cm²/hr). This net sodium transport was electroneutral and blocked by mucosal amiloride (0.5–1 mM/liter) or by interstitial ouabain (1 mM/liter). These results are consistent with the existence of a Na⁺/H⁺ exchanger on the luminal side of the bovine main pancreatic duct.     

Mechanosensory function of microvilli of the kidney proximal tubule

Normal variations in glomerular filtration induce proportional changes in proximal tubule Na+ reabsorption. This "glomerulotubular balance" derives from flow dependence of Na+ uptake across luminal cell membranes; however, the underlying physical mechanism is unknown. Our hypothesis is that flow-dependent reabsorption is an autoregulatory mechanism that is independent of neural and hormonal systems. It is signaled by the hydrodynamic torque (bending moment) on epithelial microvilli. Such signals need to be transmitted to the terminal web to modulate Na+-H+-exchange activity. To investigate this hypothesis, we examined Na+ transport and tubular diameter in response to different flow rates during the microperfusion of isolated S2 proximal tubules from mouse kidneys. The data were analyzed by using a mathematical model to estimate the microvillous torque as function of flow. In this model, increases in luminal diameter have the effect of blunting the impact of flow velocity on microvillous shear stress and, thus, microvillous torque. We found that variations in microvillous torque produce nearly identical fractional changes in Na+ reabsorption. Furthermore, the flow-dependent Na+ transport is increased by increasing luminal fluid viscosity, diminished in Na+-H+ exchanger isoform 3 knockout mice, and abolished by nontoxic disruption of the actin cytoskeleton. These data support our hypothesis that the "brush-border" microvilli serve a mechanosensory function in which fluid dynamic torque is transmitted to the actin cytoskeleton and modulates Na+ absorption in kidney proximal tubules.     

Trimethylamine-N-oxide (TMAO) increased aquaporin-2 expression in spontaneously hypertensive rats

Recent evidence suggests that elevated plasma levels of Trimethylamine-N-oxide (TMAO) can prolong the duration of elevated blood pressure in rats. The purpose of this study was to investigate the plasma TMAO level in Spontaneously Hypertensive Rats (SHR) and to explore the possible relationship between TMAO and aquaporin-2 (AQP-2) in the formation of hypertension. Twelve-week-old, male Spontaneously Hypertensive rats (SHR, n = 40) and Wistar-Kyoto rats (WKY, n = 40) were accordingly grouped into SHR group and WKY group. Each group was divided randomly into four subgroups: Untreated group, TMAO group, TMAO+Tolvaptan (TMAO+TVP) group, and TVP group, respectively. Systolic blood pressure (SBP), plasma TMAO, plasma osmolality (POsm), plasma vasopressin (PAVP), and plasma AQP-2 (PAQP-2) concentration were measured, and the expression of AQP-2 in kidney medulla was detected by RT-PCR and Western blot. At 14 weeks, rats in SHR TMAO group were shown the increased plasma TMAO, POsm, PAVP, and PAQP-2 levels, while those rats in SHR TMAO+TVP group were shown the decreased plasma TMAO, POsm, and PAQP-2 levels, but an even higher PAVP (due to the blockage of TVP to V2 receptor). These findings indicate that an increase of plasma TMAO levels in SHR leads to a higher plasma osmotic pressure, triggers the regulation of the TMAO-AVP-AQP-2 axis in SHR, elicits the greater water reabsorption, and eventually leads to hypertension.     

Cardiovascular remodelling and red blood cell Li+/Na+ countertransport in patients with type 1 diabetes and essential hypertension

Patients with type 1 (insulin-dependent) diabetes may develop a specific cardiac disease characterized by functional and structural abnormalities. The pathogenesis of the cardiac disease is poorly understood but cardiac and renal complications may coexist. Patients with overt diabetic nephropathy have increased red cell Li⁺/Na⁺ countertransport (CT), which reflects abnormal kinetic properties of the red cell membrane Na⁺/H⁺ exchange. Since the activation of Na⁺/H⁺ exchange has a key role in cell proliferation and cell growth, as well as in the regulation of cell sodium and cell pH and in the renal reabsorption of Na⁺ and bicarbonate, we have looked for relationships between red cell Li⁺/Na⁺ CT, Na⁺/H⁺ exchange and cardiovascular remodeling in patients with type 1 diabetes, essential hypertension and idiopathic familiar cardiomyopathy. In type 1 diabetes the maximal velocity of Li⁺/Na⁺ CT is positively correlated with interventricular septum thickness and the left ventricular wall to lumen ratio. Similar results were obtained in patients with essential hypertension. In these patients an increased Li⁺/Na⁺ CT is also associated with severe and drug-resistant hypertension and with significant vascular remodelling, estimated by the minimal post-ischaemic vascular resistance in the calf. Finally, Li⁺/Na⁺ CT is negatively correlated with diastolic relaxation of the left ventricle in familiar hypertrophic cardiomyopathy. From these data it appears that widespread abnormal kinetic properties of Na⁺/H⁺ exchange, estimated by increased red cell Li⁺/Na⁺ CT, may have epistatic effects on the pathogenesis of cardiac complications of type 1 diabetes and essential hypertension.     

Cell shrinkage activates Na+/H+ exchange in dog red cells by relieving inhibition of exchange by Na+ in isotonic medium

Na⁺/H⁺ exchangers (NHE) are widely distributed transporters responsible for regulation of cell volume and pH. In isotonic medium, NHE is often low or negligible, and is strongly activated by osmotic cell shrinkage. It is reported here that extracellular Na⁺ inhibits NHE in isotonic medium, and cell shrinkage stimulates NHE by relieving this inhibition. There is more than one inhibitory Na⁺ site on each transporter. Shrinkage activates NHE by decreasing the apparent affinity for Na⁺ at the inhibitory sites. Shrinkage has no effect on the apparent affinity for Na⁺ at the substrate sites for activation of NHE.     

Some Properties of Erythrocyte Na+ -K+ -ATPase in Essenial Hypertension

The activity and some allosteric properties of Na⁺ -K⁺ -ATPase in erythrocytes and their membrane preparations (ghosts) from 57 patients with essential hypertension and 36 normotensive controls were studied. To reveal enzyme activity in whole erythrocytes the cells were pretreated with detergent Tween-20. It was found that in the patient erythrocytes the Na⁺ -K⁺ -ATPase activity was 33% less as compared to the control group. Moreover, in the patient erythmcytes the sensitivity of the enzyme to high concentrations of MgCI₂ was decreased. In contrast, no analogous changes of the enzyme were revealed in the patient ghosts. It is suggested that the erythrocytes of patients with essential hypertension contain an inhibitor of Na⁺ -K⁺ -ATPase.     

Upregulation of myocardial Na+/H+ exchanger induced by chronic treatment with a selective inhibitor

Rats exposed to prolonged administration of the NHE-1 inhibitor cariporide showed enhanced activity of the exchanger in cardiac tissue, as assessed by the rise in the steady-state pHi value in the absence of bicarbonate (7.15+/-0.01 in control vs 7.49+/-0.06 and 7.41+/-0.05 in cariporide-treated for 1 or 2 months, respectively, P<0.05). In the presence of bicarbonate, the change in pHi was blunted due to a compensatory activation of acid loading pHi regulatory mechanisms. The enhancement of NHE activity disappeared after 1 week of the inhibitor withdrawal. The kinetic analysis of H+ fluxes after an acid load revealed an increased net H+ efflux (JH+) at any given pHi value and an alkaline shift of the apparent "set-point" of the exchanger (from 7.11+/-0.02 to 7.38+/-0.04,P <0.05) in treated rats. In the presence of the PKC inhibitor chelerythrine, the "set-point" of the exchanger was normalized in the cariporide-treated rats while JH+ at acidic pHi values persisted elevated. Cardiac NHE-1 mRNA levels and protein expression were increased in cariporide-treated rats. In addition to the increased protein expression after the treatment, the normalization of the augmented "set-point" by chelerythrine suggests an increased turnover rate of the units through a PKC dependent pathway. These data demonstrate that long-term treatment with the NHE-1 inhibitor cariporide enhances the antiporter activity in cardiac tissue through an increase of the number and turnover of functional units. This finding deserves further experimental and clinical evaluations to consider whether it would be advisable a gradual withdrawal of prolonged NHE inhibition to avoid an enhanced response when the exchanger is stimulated.     

PKC mediates GnRH activation of a Na/H exchanger in goldfish somatotropes

Previous results suggest that gonadotropin-releasing hormone (GnRH) stimulation of somatotropin secretion in goldfish involves activation of Na⁺/H⁺ exchange (NHE). We tested the hypothesis that GnRH alkalinizes intracellular pH (pH_i) via protein kinase C (PKC) activation of NHE. Two types of alkalinization responses were observed in identified goldfish somatotropes preloaded with the pH-sensitive dye BCECF; the rate of pH_i changes went from a neutral or slightly negative slope to either a positive or a less negative slope relative to control. Two GnRHs, the PKC-activating TPA, and dioctanoyl glycerol each caused an alkalinization in 70-90% of somatotropes. The PKC inhibitors, Bis II and Gö6976, the NHE inhibitor amiloride, or Na⁺-free solution attenuated TPA and GnRHs actions, suggesting that PKC mediates GnRH activation of NHE. Since amiloride and Na⁺-free solution caused acidification in somatotropes at rest, regulation of basal pH_i in these cells likely involves Na⁺ flux through amiloride-sensitive NHE.     

Na+/H+ exchange and its inhibition in cardiac ischemia and reperfusion

Summary The characterization of various ion transport systems has led to a better understanding of the effects, which seem to take part in the impairment of ischemic and reperfused cardiac tissue. This review discusses the role of the Na⁺/H⁺ exchange system in the pathophysiology of ischemia and reperfusion and the beneficial effects of its inhibition.At the onset of ischemia intracellular pH (pH_i) decreases due to anaerobic metabolism and ATP hydrolysis, leading to an activation of Na⁺/H⁺ exchange. This in turn increases intracellular Na⁺ (Na⁺ _i) and activates Na⁺/K⁺ ATPase, with a consecutive increase of energy consumption. Since cellular Na⁺ and Ca⁺⁺ transport are coupled by the Na⁺/Ca⁺⁺ exchange system, which depends on the Na⁺ gradient, the high Na⁺ _i leads to increased intracellular Ca⁺⁺ (Ca⁺⁺ _i). After a certain period, Na⁺/H⁺ exchange is inactivated by a decrease of extracellular pH.In case of reperfusion the acid extracellular fluid is washed out, which reactivates Na⁺/H⁺ exchange, leading to an unfavourably fast restoration of pH_i and a second time to Na⁺ and Ca⁺⁺ _i overflow.High Ca⁺⁺ _i is assumed to be one of the main reasons for ischemic and reperfusion injury, like arrhythmias, myocardial contracture, stunning and necrosis.It seems that the inhibition of Na⁺/H⁺ exchange can interrupt this process at an early phase and prevent or delay the consequences of ischemia and reperfusion as demonstrated by numerous investigators.     

Na+/H+ exchange mediates postprandial ileal water and electrolyte transport

Feeding stimulates fluid and electrolyte absorption in the small intestine. Previous studies have suggested that Na⁺/glucose cotransport is important in initiating this response in the jejunum. The purpose of this study was to determine whether Na⁺/H⁺ exchange plays a role in meal-induced absorption. Exteriorized, neurovascularly intact jejunal and ileal loops (25 cm) were constructed in dogs. Following a two-week period of postoperative recovery, the loops of awake dogs were perfused with standard buffer alone or with increasing concentrations of amiloride, a Na⁺/H⁺ exchange inhibitor. Water, sodium, and chloride fluxes were calculated following a meal using [¹⁴C]PEG as a volume marker. The meal significantly increased absorption in both the jejunum (P<0.001) and ileum (P<0.01) in those animals perfused with buffer alone. More significantly, amiloride suppressed the increased absorption seen following a meal in the ileum (P<0.001) but not the jejunum. The response in the ileum was dose dependent. These findings suggest that a major mediator of postprandial sodium and water absorption in the ileum is the Na⁺/H⁺ exchanger.Supported by NIH R29-DK-47326 (S.W.A.), R01-DK-39879 (M.J.Z.), and a VA Merit Review (D.W.M.).Portions of this work were presented at the Annual Meeting of the American Gastroenterological Association, May 1993, Boston, Massachusetts.     

The role of p38 in the regulation of Na+-Ca2+ exchanger expression in adult cardiomyocytes

The Na(+)-Ca(2+) exchanger is crucial in the regulation of [Ca(2+)](i) in the cardiac myocyte. The exchanger is upregulated in cardiac hypertrophy and failure. This upregulation can have an effect on calcium transients and possibly contribute to diastolic dysfunction and an increased risk of arrhythmias. Here we use adenovirus mediated gene expression to examine the role of p38 MAP kinase in upregulation of the exchanger in adult cardiac myocytes. We demonstrate that p38 mediates a part of the alpha-adrenergic stimulated upregulation of the Na(+)-Ca(2+) exchanger gene. Overexpression of dominant-negative p38 isoforms and activated MKK3 and MKK6 in isolated adult cardiac myocytes demonstrates that p38 activation is sufficient for NCX1 promoter upregulation and that this is mediated primarily by the p38alpha isoform. Lastly, this work demonstrates that the p38alpha stimulated upregulation of the NCX1 promoter is mediated via the -80 CArG box element. This is the first time that a specific role for p38alpha in gene regulation has been demonstrated in isolated adult cardiomyocytes and provides an important clue to our understanding some of the factors regulating exchanger gene expression in the hypertrophic and failing heart.     

The Effect of Melittin on Na+ and Rb+ Transport in Cultured Skin Fibroblasts of the Spontaneously Hypertensive Rat

Melittin effect on transport of Na⁺ and Rb⁺(K⁺ analog) was examined in cultured skin fibroblasts originating from the Spontaneously Hypertensive, Wistar Kyoto and Wistar rats. Melittin increased both Na⁺ (²²Na⁺₊) uptake and 86Rb⁺ efflux as well as the activity of the Na⁺-pump (ouabain sensitive ⁸⁶Rb⁺ uptake) in all three preparations. The effect of the toxin was maximal at a dose of 160–240ng/10⁵ cells/ml. At this does, fibroblasts of the Spontaneously Hypertensive rat demonstrated the greatest response to melittin with respect to the increase in Na⁺ and Rb⁺ fluxes and increase in the intracellular Na⁺ concentrations. It is concluded that melittin can be utilized as a probe to delineate subtle differences in the cellular regulation of Na⁺ and K⁺ in the Spontaneously Hypertensive rat as compared with its normotensive controls.