Shiv C. Kapoor

Potassium Depletion Exacerbates Essential Hypertension

OBJECTIVE To determine the effect of potassium depletion on blood pressure in patients with essential hypertension. DESIGN Double-blind, randomized, crossover study, with each patient serving as his or her own control. SETTING Clinical research center at a university hospital. PATIENTS Twelve patients with hypertension. INTERVENTIONS Patients were placed on 10-day isocaloric diets providing a daily potassium intake of either 16 mmol or 96 mmol. The intake of sodium (120 mmol/d) and other minerals was kept constant. On day 11 each patient received a 2-litre isotonic saline infusion over 4 hours. MEASUREMENTS Blood pressure; urinary excretion rates for sodium, potassium, calcium, and phosphorous; glomerular filtration rate; renal plasma flow; and plasma levels of vasoactive hormones. MAIN RESULTS With low potassium intake, systolic blood pressure increased (P = 0.01) by 7 mm Hg (95% CI, 3 mm Hg to 11 mm Hg) and diastolic pressure increased (P = 0.04) by 6 mm Hg (CI, 1 mm Hg to 11 mm Hg), whereas plasma potassium concentration decreased (P less than 0.001) by 0.8 mmol/L (CI, 0.4 to 1.0 mmol/L). In response to a 2-litre isotonic saline infusion, the mean arterial pressure increased similarly on both diets but reached higher levels on low potassium intake (115 +/- 2 mm Hg compared with 109 +/- 2 mm Hg, P = 0.03). Potassium depletion was associated with a decrease in sodium excretion (83 +/- 6 mmol/d compared with 110 +/- 5 mmol/d, P less than 0.001). Plasma renin activity and plasma aldosterone concentrations also decreased in patients during low potassium intake, but concentrations of arginine vasopressin and atrial natriuretic peptide, glomerular filtration rate, and renal plasma flow were unchanged. Further, low potassium intake increased urinary excretion of calcium and phosphorus and of plasma immunoreactive parathyroid hormone levels. CONCLUSION Dietary potassium restriction increases blood pressure in patients with essential hypertension. Both sodium retention and calcium depletion may contribute to the increase in blood pressure during potassium depletion.

Natriuretic Effect of Calcium-Channel Blockers in Hypertensives

This double-blind, randomized, crossover trial characterizes the acute natriuretic response to calcium-channel blockers (CCB) and investigates the role of hemodynamic and hormonal factors in mediating the natriuresis. Thirteen male subjects with essential hypertension received a single oral 20-mg dose of nifedipine or 120 mg of diltiazem. Renal functional and hemodynamic measurements were performed prior to and hourly for 4 hours following medication. Subjects then received these medications for 4 weeks at which time the above studies were repeated. Urinary sodium excretion increased within 60 minutes of CCB administration and the natriuresis was sustained for 4 hours. Cumulative sodium loss during the 4 hours of study was greater with nifedipine (43 +/- 12 mmol) than with diltiazem (18 +/- 6 mmol) (P less than 0.05). Despite natriuresis, urinary potassium excretion was decreased by both agents. Even though both drugs decreased the mean arterial pressure, inulin and paraaminohippurate (PAH) clearances were not altered. Plasma aldosterone concentrations decreased, plasma catecholamine concentrations increased, whereas plasma-renin activity was unchanged with both drugs. Body weight, glomerular filtration rate (GFR), renal plasma flow, plasma-renin activity, plasma aldosterone, and catecholamine concentrations were unchanged following 4 weeks of therapy. The acute natriuretic response after 4 weeks of therapy was similar to the response noted after the first dose. This study concludes that CCB are acutely natriuretic. Despite systemic hypotension, renal hemodynamics are unaltered during CCB therapy. Suppression of aldosterone as well as direct tubular effects of these drugs may mediate the natriuresis. Chronic therapy with CCB does not modify the acute natriuretic response to these agents.

Effects of Pb2+ added in vitro on Ca2+ movements in isolated mitochondria and slices of rat kidney cortex

We have studied the effects of Pb2+ added in vitro on the movements of Ca2+ in renal cortical mitochondria and tissue slices. The isolated mitochondria rapidly accumulated 45Ca2+ at 25 degrees by a mechanism that was dependent on respiration and inhibited 96% by ruthenium red. A concentration of 10 microM Pb2+ inhibited the Ca2+ accumulation at least as effectively as did ruthenium red. About 20% of the Ca2+ accumulation persisted at 1 degrees with a similar sensitivity to inhibitors, including 60% inhibition by Pb2+. Similar results were obtained when the accumulation of Ca2+ at 25 degrees was measured by means of a calcium-sensitive electrode, Pb2+ inhibiting by 80%. Calcium that had been accumulated by mitochondria at 25 degrees was released completely by the ionophore A23187 or by 10 microM Pb2+. The release induced by Pb2+ was greatly inhibited by ruthenium red. The Ca2+ content of tissue slices of renal cortex increased 4-fold during incubation at 1 degree while the Ca2+ content of mitochondria within the slices more than doubled, the latter being determined by isolation of mitochondria from the slices after incubation. The presence of Pb2+ (200 microM) in the incubation medium of the slices substantially reduced the entry of Ca2+ into the whole slices and into mitochondria within the slices. When the slices preincubated at 1 degree were warmed to 25 degrees in oxygenated medium, they brought about a net extrusion of Ca2+, some of which was derived from the mitochondria; Pb2+ did not alter the final level of Ca2+ then attained in the slices, but it caused a significant decrease in the quantity retained in the mitochondria. We conclude that Pb2+ both inhibits the uptake of Ca2+ by renal cortical mitochondria and displaces Ca2+ from them, these effects occurring whether the mitochondria are isolated or in situ.

PRESERVATION OF RENAL RESERVE IN CHRONIC RENAL DISEASE

Protein-induced increases in glomerular filtration rate (GFR), termed renal reserve, is said to be abrogated with the onset of renal disease. However, this notion is inconsistent with the results from animal studies which suggest that alterations in protein intake modulate the glomerular hemodynamics in experimental renal disease. Accordingly, 12 normal subjects and 15 patients with renal disease received a protein meal providing 1 g/kg body weight protein. The subjects were pretreated with either placebo or an angiotensin I converting enzyme inhibitor, enalapril. A significant (P less than 0.05) increase in inulin and para-aminohippurate (PAH) clearance was noted in normal subjects as well as in patients with renal disease. The increase in GFR over basal values in normal subjects (28 +/- 9%), patients with moderate renal failure (20 +/- 13%), and advanced renal failure (21 +/- 14%) was not different. Plasma renin activity was unchanged following protein meal in the placebo studies although it increased following enalapril administration. Enalapril pretreatment did not alter the glomerular vasodilation and hyperfiltration following protein meal. We conclude that protein meal induces glomerular hyperfiltration in renal disease and that this protein-induced hyperfiltration is not mediated by angiotensin II. Because glomerular hyperfiltration is implicated in the progression of renal disease, these data suggest that even in patients who have advanced renal failure, high-protein diets may exert a detrimental effect on the kidney.

Morphological and physiological studies of rat kidney cortex slices undergoing isosmotic swelling and its reversal: a possible mechanism for ouabain-resistant control of cell volume

SummarySlices of rat kidney cortex were induced to swell by preincubation at 1°C in an isotonic Ringers solution, and their capacity to reverse swelling, by net extrusion of cellular water, was studied during subsequent incubation at 25°C. The recovery from swelling was prevented by the respiratory inhibitor, antimycin A. On the other hand, extrusion of water was little affected by ouabain. The extrusion of water continuing in the presence of ouabain (but not that in its absence) was significantly reduced when furosemide was added or when medium Cl− was replaced by NO3−, or I−. There was substantial variability in the morphological appearance of cells within the cortical slices. Different segments of the nephron showed different structural changes during swelling and its reversal, the proximal tubules being most markedly affected. Proximal tubular cells of swollen slices showed disorganization of brush borders and expansion of their apical surfaces, and contained vesicles in their apical cytoplasm. Upon recovery at 25°C, the apical portions of these cells showed reversal of the expansion, but some apical vesicles remained. These vesicles were much more numerous after recovery in the presence of ouabain, but they were much reduced in numbers, or totally absent, when recovery took place in the presence of furosemide or absence of Cl−, with or without ouabain. The vesicles seen in the presence of ouabain alone appeared to fuse with each other and with infoldings of the basolateral plasma membrane. Rather similar results were obtained with distal tubular cells in the slices. We suggest that volume regulation in the proximal and distal tubular cells proceeds by way of two mechanisms. The first consists of extrusion of water coupled to the ouabain-sensitive transport of Na+ and K+. The other proceeds by way of an ouabain-resistant entry of water into apical cytoplasmic vesicles, following furosemide-sensitive movements of Cl− and Na+; the vesicles then expel their contents by exocytosis at the basolateral cell borders.

Uptake of inorganic lead in vitro by isolated mitochondria and tissue slices of rat renal cortex.

Slices of rat renal cortex were shown to take up Pb2+ during incubation in vitro; Pb2+ was also shown to enter mitochondria within the slices. The uptake of Pb2+ by isolated mitochondria was inhibited by N-3, La3+ and ruthenium red. A steady state of uptake was attained within 60 sec. The concentration dependence of uptake was complex; maximum uptake was attained at 25 microM and inhibition ensued at higher concentrations. A substantial inhibitor-resistant component of Pb2+ uptake was noted, especially at medium Pb2+ concentrations greater than 25 microM, and these concentrations also inhibited respiration state 3. The effects on respiration were reduced if the mitochondria had been preincubated with ruthenium red. Slices of renal cortex incubated at 1 degree in medium with various concentrations of Pb2+ showed two fractions of uptake, one saturating at 50-100 microM external Pb2+ and the other at 150-200 microM. Subsequent incubation for 60 min at 25 degrees led to further uptake at all concentrations. Upon isolation of mitochondria from incubated slices, significant amounts of Pb2+ were detected in the mitochondria within 5 min of addition of Pb2+ (200 microM), with maximum attained at 30 min. Electron microscopy of slices showed electron-dense particles, apparently of Pb2+, in the cortical cells but the greatest concentration was deposited in the basement membranes. The results indicate the importance of the basement membrane in limiting access of Pb2+ to cortical cells, and of mitochondria in accumulating Pb2+ once it is in the cells. They also illustrate the importance of interactions between Pb2+ and Ca2+.

Inhibition of mitochondrial oxidative metabolism by SKF-525a in intact cells and isolated mitochondria

We have examined the effects of various concentrations of SKF-525A (beta-diethylaminoethyldiphenylpropyl acetate X HCl) on the energy metabolism of liver slices, isolated liver mitochondria, and two types of ascites tumor cells, as well as on ion transport in liver slices. In liver slices, 0.2 to 1.0 mM SKF-525A caused an initial stimulation of O2 uptake which was followed, at 0.5 to 1.0 mM, by a progressive inhibition of O2 consumption, a fall of slice ATP content, and a reduced transport of K+, Na+ and Ca2+. In isolated mitochondria, we studied the effects of SKF-525A on the rate of respiration and on the oxidation-reduction responses of NAD(P)+ and cytochrome b in the presence of various substrates. The results suggest that SKF-525A had three distinct actions on liver mitochondria, viz. an uncoupling action at low concentrations (0.02 to 0.17 mM); at higher concentrations (0.2 to 0.5 mM) an inhibition of the oxidation of NAD(P)+-linked substrates, exerted close to the substrate level; also at 0.2 to 0.5 mM, a less effective inhibition of electron transfer at a point between cytochrome b and O2 in the electron-transfer chain. Experiments on O2 consumption and cytochrome b oxidation-reduction changes in ascites cells showed only the first two of these effects in the intact tumor cells. We conclude that inhibition of mitochondrial energy-conserving reactions by SKF-525A can have a marked influence on energy-requiring aspects of liver-cell metabolism, one example of which is inhibition of cation active transport.

Effects of inorganic lead in vitro on ion exchanges and respiratory metabolism of rat kidney cortex.

The effects of Pb2+ added in vitro to tissue slices, isolated tubules and isolated mitochondria of rat kidney cortex have been studied. Slices were depleted of K+ and loaded with Na+, Cl− and water by pre-incubation at 1° C, and reversal of these changes was then induced by incubation under metabolically favourable conditions. The net reaccumulation of K+ was reduced by a maximum of 30% when Pb2+ was present in the medium, the maximal effect being caused by 200 μM Pb2+. Lead also caused a reduction of Na+ extrusion which was approximately equimolar with its effect on K+, but it did not affect the extrusion of Cl− and water. The initial rates of the net, active movements of K+ and Na+ were not altered by Pb2+, divergence from control values only being noted after 15–30 min incubation. The O2 consumption and the ATP content were 25–30% lower in slices incubated with 200 μM Pb+ than in control slices; the effect on ATP content was not observed until incubation had continued for 30 min. In tubules isolated from the renal cortex, the rate of respiration (50%) and ATP content (30%) were also partly reduced by 200 μM Pb2+. The consumption of O2 by mitochondria isolated from the cortex was much more sensitive to Pb2+ added in vitro than the respiration of intact cells; the rate of respiration in state 3 (presence of phosphate acceptor) and the respiratory control ratio were drastically reduced, with half-maximal inhibition at 30 and 20 μM Pb2+ respectively. Comparison of the effects of Pb2+ on energy metabolism and ion transport of the slices with the corresponding effects of antimycin A and ouabain suggests that Pb2+ inhibited K+ and Na+ transport mainly as a consequence of a primary inhibition of the provision of ATP.

Effect of Dietary Protein on Glomerular Renin Secretion

Previous studies from our and other laboratories demonstrated that dietary protein restriction lowers plasma renin activity by impairing renin release. The effect of protein intake on glomerular renin secretion has not been investigated. Accordingly, we studied male Sprague-Dawley rats weighing 180 to 200 g for 3 weeks that were receiving isocaloric diets that provided either standard 20% protein (SP) or low 6% protein (LP). Renin secretion was measured in the glomeruli isolated from these rats, at baseline and following stimulation with arachidonic acid and isoproterenol. The activity of plasma renin (3.0 +/- 0.5 ng/mL/min on SP v 1.1 +/- 0.1 ng/mL/min on LP) was significantly (P < 0.02) lower on LP intake. In contrast, glomerular renin content (22.9 +/- 0.7 ng/micrograms protein on SP v 32.3 +/- 1.4 ng/micrograms protein on LP) was significantly (P < 0.01) higher on the LP diet. Furthermore, renin secretion (ng/mL/h) from the isolated glomeruli at baseline (3.9 +/- 1.0 on SP v 12.5 +/- 3.0 on LP, P < 0.02), and following incubation with arachidonic acid 10(-5) mol/L (5.9 +/- 1.7 on SP v 19.6 +/- 3.1 on LP, P < 0.005), and isoproterenol 10(-3) mol/L (6.0 +/- 0.5 on SP v 17.3 +/- 3.3 on LP, P < 0.01) was significantly higher on the LP diet. These studies suggest that dietary protein restriction impairs in vivo renin release. In contrast, in vitro glomerular renin release is augmented by protein restriction. The factors modulating in vivo renin release require further characterization.