A B S T R A C T The major renal adaptive changes in response to selective dietary phosphate restriction are a marked reduction in urinary excretion of phosphate and an increased urinary excretion of calcium; at the cellular level, there is selective increase in renal cortical brush border membrane phosphate uptake and increase in specific activity of alkaline phosphatase. In the present study we examined whether these functional and biochemical adaptive changes could be blocked by drugs known to inhibit protein synthesis.Administration of actinomycin D or cycloheximide to rats switched from a diet with normal phosphate content (0.7%) to a diet with low (0.07%) phosphate content either completely (actinomycin D) or partially (cycloheximide) prevented the expected decrease in urinary excretion of phosphate and increase in the urinary excretion of calcium. The specific activity of alkaline phosphatase measured in crude membrane fraction (washed 100,000 g pellet) from renal cortical homogenate in animals fed a low phosphate diet and treated with actinomycin D or with cycloheximide was significantly lower than in control animals also on a low phosphate diet receiving placebo; but there were no differences between treated and untreated animals in the activities of two other brush border enzymes, y-glutamyltransferase and leucine aminopeptidase. Actinomycin D administered to rats maintained on a normal phosphate diet throughout the course of the experiment caused an increase in the urinary excretion of phosphate on the last (6th) day of the experiment but did not change urinary excretion of calcium. In acute clearance experiments, infusion of actinomycin D to rats adapted to a low phosphate diet did not increase fractional excretion of phosphate.In separate experiments, using the same dietary protocol as above, brush border membrane fraction (vesicles) was prepared from renal cortex of rats sacrificed at the end of the experiment. In this preparation Na+-dependent 32Pi and D-[3H]glucose uptake and activities of brush border enzymes membrane were determined. Brush border membrane vesicles prepared from rats fed a low phosphate diet showed significantly higher Na+-dependent 32Pi uptake compared with rats fed a normal phosphate diet. This increase in -'Pi uptake was completely prevented when rats on a low phosphate diet were simultaneously treated with actinomycin D. These differences were specific for 32Pi transport as no differences were observed in D_[3H]glucose uptake among the three groups. There was a positive correlation (r = 0.82, P < 0.01) between 32Pi uptake and specific activity of alkaline phosphatase measured in aliquots ofthe same brush border membranes, whereas no such correlation was observed with two other brush border membrane enzymes y-glutamyltransferase and leucine aminopeptidase.These observations show that actinomycin D prevents both the functional and cellular renal adaptive changes induced by a low phosphate diet. Taken together, these observations suggest that renal adaptation to a low phosphate ...

A B S T R A C T Because glomerular functions are modulated by numerous humoral agents, probably acting through cyclic nucleotides, the effects of some polypeptide hormones and biogenic amines on cyclic AMP (cAMP) and cyclic 3',5'-guanosine monophosphate (cGMP) were studied in glomeruli isolated from rat renal cortex. Glomeruli and cortical tubules were prepared by a combination of sieving and density-gradient centrifugation. Under basal conditions, the contents of cAMP and cGMP in glomeruli were significantly higher than in tubules and unfractionated renal cortical tissue.Histamine caused a striking increase in cAMP in glomeruli (+A% 675±87) and, to a lesser degree, increased cAMP in tubules (+A% 103+25) or in tissue slices. This stimulation was dose-dependent in the range of 1 ,uM-1 mM histamine. Metiamide (an H2-antagonist), but not pyrilamine (an H,-antagonist) blocked the effect of histamine on cAMP, which indicates that histamine causes its effect via interaction with H2 receptors. Histamine caused less extensive increases in cGMP in both glomeruli and tubules. Carbamylcholine caused a marked increase in cGMP in glomeruli (+A 295±7) and a much lower increase in tubules (+A% 70+20); these effects were blocked by 1334atropine. Parathyroid hormone (1 ug/ml) increased cAMP and, to a much lesser degree, also cGMP in glomeruli. In tubules, parathyroid hormone caused nmuch more extensive increases in cAMP than in glomeruli; no changes, or rather a small decline in cGMP, was observed. Angiotensin-I1 (2 uM) markedly lowered cAMP in glomeruli (-A% -45±8) and in tubules (-A% 33±7) but had no effect on cGMP. Bradykinin (20 ,M) did not consistently influience either cAMP or cGMP in glomeruli or tubules.Present results demonstrate that cAMP and cGMP metabolism in glomeruli are controlled independently by humoral agents known to alter glomerular functions in vivo. Our findings are consistent with the view that histamine and cholinergic agents generated and (or) released locally in glomeruli or in their vicinity may play important roles as mediators of immunopathological injury of glomeruli, and that these effects are mediatecl by cAMP and (or) cGMP through interaction with H2 receptors and muscarinic receptors. Likewise, our results suggest that the effects of angiotensin-II and parathyroid hormone on glomerular dynamics may be mediated by cyclic nucleotides.Thus, we surmise that extrarenal as well as intrarenal humoral agents may play an important role in the pathology and physiology of glomeruli through mediation of either cAMP, cGMP, or both. INTRODUCTIONIt has become increasingly evident in recent years that many extrarenal and intrarenal humoral agents mraymodulate diverse aspects of glomerular function (1). For example, the administration of agents such as para- thyroid hormone (PTH)' (2), angiotensin (3, 4), bradykinin, acetylcholine (5,6), and others (1) causes changes in glomerular microcirculation, hydraulic permeability, or in the ultrafiltration properties of glomerular membrane. Perhaps more importantly,...

Previous findings suggest that alkaline phosphatase (Alk Pase) may be involved in phosphate transport. Since phosphate reabsorption is enhanced in the kidney and duodenum of animals stabilized on a low-phosphorus diet (LPD), Alk Pase was measured in the kidney, small intestine, and other tissues in LPD rats. In particulate fractions from the renal cortex, intestine, renal medulla, liver, and heart ventricle from LPD rats the activity of Alk Pase was significantly increased but the activities of other plasma membrane enzymes were not different between control and LPD groups. The increased Alk Pase in the renal cortex was localized to the brush border of the proximal tubule histochemically and by measurement of Alk Pase in brush-border preparations. Also in the renal cortex, typical enzymes associated with mitochondria, lysosomes, and cytosol were unchanged with the exception of cytosolic adenosine 3',5' cyclic-monophosphate phosphodiesterase, which was increased in LPD rats. Alk Pase in the renal cortex and intestine may play a role in the enhanced phosphate reabsorption in LPD animals.

Experiments were performed to determine whether longitudinal and circular muscles from various regions of stomach and small bowel had the capacity to convert arachidonic acid (AA) to prostaglandins (PGs). PG production by the microsomal fractions of isolated muscles was assayed by determining the conversion of [14C]AA to 14C-labeled 6-keto-PGF1 alpha, PGF2 alpha, PGE2, PGD2, PGA2, and thromboxane B2. Individual PGs were identified by thin-layer chromatography. The metabolism of [14C]AA to [14C]PGs was linearly related to substrate concentration, enzyme concentration, and incubation time at 37 degrees C and was inhibited in a dose-dependent manner by indomethacin. Longitudinal and circular muscles from all tested regions (corpus, fundus, antrum, pylorus, duodenum, jejunum, and ileum) synthesized PGs. In all regions the major end products of AA metabolism were 6-keto-PGF1 alpha, PGE2, and PGF2 alpha. The data indicate that circular and longitudinal muscles from all regions of the stomach and small bowel contain the enzymatic apparatus necessary to convert AA into prostaglandins.

Serotonin (5-hydroxytryptamine) is known to influence glomerular function and may have an important role in the pathogenesis of glomerulopathies. Because serotonin acts in nonrenal tissues through mediation of cyclic nucleotides, we investigated in vitro its effect on cAMP and cyclic guanosine monophasphate (cGMP) in tissue slices and isolated glomeruli from rat kidney. Serotonin increased cAMP 161 +/- 35% but not cGMP in renal cortex; it had no effect on cyclic nucleotides in medulla and papilla. In isolated glomeruli, serotonin elicited a dose-dependent (in the range of 10-7 to 10-4 M) increase in cAMP; the maximum increase over basal values was 376 +/- 45%. Serotonin increased cAMP either in the presence or in the absence of a cAMP phosphodiesterase inhibitor. In tubular fraction, serotonin elevated cAMP to a much lesser degree (82 +/- 15%). Neither in glomeruli nor in tubules did cGMP concentrations change in response to serotonin, but carbamylcholine, a known cGMP agonist, significantly increased cGMP concentrations. The increase in cAMP in response to serotonin was blocked (greater than 85% inhibition) by equimolar concentrations of serotonin antagonists methysergide and cinanserine. Results of this study demonstrate that interaction of serotonin with receptors in the kidney, particularly in the glomeruli, cause a striking increase in cAMP concentrations without detectable changes in cGMP concentrations. These findings suggest that serotonin, either synthesized in the kidney or released locally from platelets aggregated in glomeruli (for example, in association with immunopathologic injury) may exert of modulate its physiologic or pathologic effects via mediation of cAMP.