Calcium absorption or secretion by rat ileum in vitro: effects of dietary calcium intake.
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Animals↗
Biomedical subjects
D V Kimberg
Publications and source records attributed to D V Kimberg.
Effects of prostaglandins and cholera enterotoxin on intestinal mucosal cyclic AMP accumulation. Evidence against an essential role for prostaglandins in the action of toxin.
Both cholera enterotoxin and certain prostaglandins have been shown to stimulate intestinal fluid secretion in vivo, to cause ion flux changes in vitro similar to those caused by addition of cyclic 3',5'-adenosine monophosphate (cyclic AMP), and to activate intestinal mucosal adenyl cyclase. It has been suggested that the effects of the enterotoxin on intestinal cyclic AMP metabolism may be indirect, and that locally synthesized prostaglandins may serve as required intermediates for the effects of the enterotoxin in activating intestinal mucosal adenyl cyclase. In order to clarify certain aspects of the mechanisms by which these two agents alter intestinal mucosal cyclic AMP metabolism and ion transport, their effects on cyclic AMP accumulation in rabbit ileal mucosa were examined in vitro. Addition of 5 mug per ml (75 mug per 150 mg mucosa) of purified cholera enterotoxin produced a peak increase in cyclic AMP level in 3 h but there was a time delay of at least 30 min before any effect was observed. Inhibition of cyclic nucleotide phosphodiesterase with theophylline failed to reduce this time delay. In contrast, addition of prostaglandin E(1) (PGE(1)) increased the cyclic AMP level rapidly, a peak effect being observed in 2 min. The time of the peak prostaglandin-induced changes in cyclic AMP level and short-circuit current correlated closely. A maximal increment in cyclic AMP level was achieved with 5 x 10(-5) M PGE(1). When 10(-4) M PGE(1) was added to mucosa already maximally stimulated with cholera toxin, the resulting cyclic AMP level was equal to the sum of the levels reached when each agent was added alone. Furthermore, the effects of the enterotoxin on mucosal cyclic AMP levels were not influenced by indomethacin under conditions where mucosal prostaglandins synthesis was inhibited. The results suggest that endogenous prostaglandins do not provide an essential link in the activation of intestinal mucosal adenyl cyclase by cholera enterotoxin. The present study also indicates that the effect of cholera enterotoxin on intestinal mucosal cyclic AMP metabolism involves a definite time delay which is not due to cyclic nucleotide phosphodiesterase activity.
Adenylyl Cyclases↗
Effects of dietary calcium restriction and chronic thyroparathyroidectomy on the metabolism of (3H)25-hydroxyvitamin D3 and the active transport of calcium by rat intestine.
Previous studies have shown that chronically thyroparathyroidectomized (TPTX) rats, fed a diet with restricted calcium but adequate phosphorus and vitamin D content, have higher levels of intestinal calcium absorption than controls. The results of recent acute experiments have suggested that parathyroid hormone (PTH) may be essential for regulating the renal conversion of 25-hydroxyvitamin D(3) (25-OH-D(3)) to 1,25-dihydroxyvitamin D(3) [1,25-(OH)(2)-D(3)] in response to dietary calcium deprivation. Since 1,25-(OH)(2)-D(3) is the form of the vitamin thought to be active in the intestine, increases in calcium transport mediated by this metabolite would not be expected to occur in the absence of the parathyroid glands if the preceding model is correct. The present study was undertaken to examine the chronic effects of both dietary calcium restriction and the absence of PTH on the metabolism of [(3)H]25-OH-D(3) and duodenal calcium-active transport in rats given thyroid replacement. These relatively long term studies confirm earlier observations which indicated that the adaptation of calcium absorption to a low calcium intake occurs in both sham-operated and TPTX animals. The present studies also demonstrated that despite reduced levels of 1,25-(OH)(2)-D(3) in the plasma of chronically TPTX animals fed a low calcium diet, the accumulation of this metabolite in at least one target tissue, intestinal mucosa, is identical in both the sham-operated and TPTX groups. A reduced, but continued level of 1,25-(OH)(2)-D(3) production, together with its selective accumulation by intestinal mucosa, probably explains the calcium adaptation which is observed inspite of the chronic absence of the parathyroid glands.
Animals↗
Vasoactive intestinal peptide stimulation of adenylate cyclase and active electrolyte secretion in intestinal mucosa.
Vasoactive intestinal peptide (VIP), originally isolated from hog small intestinal mucosa, has been shown to cause small intestinal secretion. More recently, this peptide has been identified in the plasma and tumors of patients with the so-called "pancreatic cholera" syndrome. In order to explore the possible role of VIP in the pathogenesis of this syndrome, we examined the effects of this peptide and other hormones on the cyclic AMP levels, adenylate cyclase activity, and ion transport in in vitro preparations of ileal mucosa. In rabbit ileal mucosa, VIP (20 mug/ml) caused a prompt fivefold increase in cyclic AMP level, whereas nine other hormones, which have been postulated to cause intestinal secretion, failed to exert such an effect. Pentagastrin and glucagon also failed to increase cyclic AMP levels in canine ileal mucosa. An increase in mucosal cyclic AMP levels was observed at a VIP concentration of 0.1 mug/ml and appeared to be nearly maximal at 2.0 mug/ml. VIP (100 mug/ml) stimulated adenylate cyclase activity in a membrane preparation from rabbit ileal mucosa. Secretin (6.0 x 10(-5) M) failed to do so. When added to the serosal side of isolated rabbit ileal mucosa clamped in an Ussing chamber, VIP (2 mug/ml) increased short-circuit current (SCC) and caused net secretion of both Cl and Na. Net Cl secretion exceeded net Na secretion. These effects of VIP on mucosal cyclic AMP metabolism and ion transport are similar to those observed with cholera enterotoxin and certain prostaglandins. VIP was also tested with normal human ileal mucosa. At a concentration of 2 mug/ml it caused a fivefold increase in cyclic AMP level and an increase in SCC of the same magnitude as that caused by 5 mM theophylline. Addition of a second 2-mug/ml dose of VIP and addition of theophylline after VIP produced no further change in SCC. We conclude the VIP stimulates adenylate cyclase and active ion secretion in both rabbit and human ileal mucosa. This may be related to the pathogenesis of diarrhea in patients with the pancreatic cholera syndrome.
Adenylyl Cyclases↗
Active secretion of calcium by adult rat ileum and jejunum in vitro.
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Age Factors↗
Effects of cortisone administration on the metabolism and localization of 25-hydroxycholecalciferol in the rat.
Glucocorticoid administration is known to decrease calcium absorption in vivo and the vitamin D-dependent active transport of calcium by rat duodenum in vitro. The basis for this antivitamin D-like effect of glucocorticoids is unclear. Previous studies in the rat failed to demonstrate an effect of glucocorticoid treatment on the hepatic conversion of the parent vitamin to 25-hydroxycholecalciferol (25-HCC). Moreover, pharmacologic doses of 25-HCC did not restore intestinal calcium transport to normal. The results of these experiments suggested that if indeed glucocorticoids interfere with the metabolism of vitamin D, the step involved must be subsequent to 25-hydroxylation. The present studies demonstrate that the administration of cortisone to vitamin D-deficient rats does not affect the rate of conversion of a physiologic dose of [(3)H]25-HCC to the biologically important metabolite, 1,25-dihydroxycholecalciferol (1,25-DHCC). Furthermore, pretreatment with glucocorticoids affects neither the tissue distribution nor the subcellular localization on or in intestinal mucosal cell nuclei of 1,25-DHCC. Of note is the fact that 1,25-DHCC is currently considered to be the "tissue-active" form of the vitamin in the intestine. Whereas tissues from cortisone-treated animals had increased concentrations of the biologically less active 24,25-DHCC, the physiologic significance of this observation remains unclear. The results of the present studies strongly support the concept that the antivitamin D-like effects of glucocorticoids in the intestine are due to hormonal influences on the biochemical reactions responsible for calcium transport. While the effects of these hormones are opposite in direction to those of vitamin D, they occur by a mechanism that is independent of a direct interaction with either the vitamin or its biologically active metabolites.
Animals↗
Effects of cycloheximide on the response of intestinal mucosa to cholera enterotoxin.
Prior studies have indicated that effects of cholera enterotoxin (CT) on the small intestine are delayed in onset and involve an interaction with adenyl cyclase in the mucosa. It has also been shown that the administration of cycloheximide to rabbits in doses which inhibit crypt cell mitoses (20 mg/kg), diminishes CT-induced fluid production in jejunal loops. These latter studies have been interpreted as indications that CT-related intestinal secretion is a crypt cell function and that it is mediated by a CT-induced protein. The present study was undertaken to delineate more precisely the nature of the interaction in the intestine between cycloheximide and cholera toxin. Pretreatment of rabbits with cycloheximide reduced by 60% the secretory response to CT in isolated ileal loops with intact blood supply. Sodium and chloride flux measurements on mucosa isolated from these and control loops indicated that this antisecretory effect of cycloheximide persists in vitro. Measurements of radioactive leucine incorporation into mucosal protein indicated that the dose of cycloheximide employed inhibited protein synthesis by 90%. This inhibitory effect was shown to be independent of any effect of cycloheximide on amino acid uptake across the brush border. Measurements of adenyl cyclase activity and cyclic AMP levels in ileal mucosa of cycloheximide pretreated and control animals indicated that cycloheximide did not diminish the CT-induced increases in these parameters. These observations demonstrate that cycloheximide reduces CT-induced intestinal fluid production without interfering with the CT-induced augmentation of adenyl cyclase activity or the consequent rise in cyclic. AMP concentration. Since the antisecretory effect of cycloheximide persists in vitro, it probably involves a direct interaction of the antibiotic with mucosal cell ion transport mechanisms rather than an indirect effect mediated by other humoral or neurogenic factors. The present observations also suggest that the secretory response of the intestine to CT involves neither the synthesis of new adenyl cyclase nor that of a protein modifying its activity.
Adenylyl Cyclases↗
Effects of 1,25-dihydroxycholecalciferol on intestinal calcium transport in cortisone-treated rats.
The administration of glucocorticoids may decrease intestinal calcium absorption in vivo and the active transport of calcium in rat duodenum in vitro. It has been suggested that this apparent "anti-vitamin D-like" effect of steroid hormones may be related to alterations in vitamin D metabolism. In order to test this hypothesis, vitamin D-deficient control and cortisone-treated rats were given an intraperitoneal injection of 5.5 IU of 1,25-dihydroxycholecalciferol (1,25-DHCC), the probable end-organ active vitamin D metabolite in the intestine, and 16 h later studies of duodenal calcium transport were performed in modified Ussing chambers. In the vitamin D-deficient state, cortisone administration was associated with a diminution in J(MS), J(Net), and the flux ratio (J(MS)/J(SM)). While the magnitude of the increases in J(MS) and J(Net) that resulted from 1,25-DHCC treatment were approximately the same in control and cortisone-treated animals, 1,25-DHCC failed to restore these parameters to "normal levels" in the steroid-treated rats. Furthermore, contrary to the results obtained in the saline-treated controls, 1,25-DHCC failed to reduce J(SM) in the duodenum from cortisone-treated rats. The cortisone-related defect in calcium transport was due to alterations in both unidirectional calcium fluxes (decrease in J(MS) and increase in J(SM)), such that the J(Net) and the flux ratio (J(MS)/J(SM)) were only approximately 50% of the levels achieved in vitamin D-deficient control animals repleted with the same dose of 1,25-DHCC. The administration of 1,25-DHCC was accompanied by a marked increase in the serum calcium levels of control rats, but there was no such response in the cortisone-treated group. The results support the concept that under the conditions of these experiments in the rat the apparent antagonism between glucocorticoids and vitamin D may be due to steroid hormone-related alterations in end organ function that are independent of any direct interaction between the hormone and the vitamin and that cannot be reversed by the vitamin.
Animals↗
Alkaline phosphatase. Possible induction by cyclic AMP after cholera enterotoxin administration.
The present studies were undertaken to determine the role, if any, of cyclic 3',5'-adenosine monophosphate (cyclic AMP) as a chemical inducer of rat liver alkaline phosphatase. Cholera enterotoxin, given intravenously to rats, led to a rapid rise in the activity of hepatic adenyl cyclase that was 7(1/2) times greater than control values in 6 h. Cyclic AMP levels were also significantly increased above control values while the activity of cyclic nucleotide phosphodiesterase was unchanged. Hepatic alkaline phosphatase activity was increased 5(1/2) times above control in 12 h, but its rise followed that of adenyl cyclase and cyclic AMP by several hours. Cycloheximide inhibited the rise of hepatic alkaline phosphatase but not that of adenyl cyclase. The administration of glucagon, a known stimulator of hepatic adenyl cyclase, and of dibutyryl cyclic AMP, led to similar striking increases in hepatic alkaline phosphatase activity. This alkaline phosphatase increase was blocked by the prior administration of cycloheximide. Bile duct ligation, a known stimulator of hepatic alkaline phosphatase activity, failed to produce any significant changes in adenyl cyclase or cyclic AMP. Concomitant treatment of rats with bile duct ligation and cholera enterotoxin or bile duct ligation and glucagon, had no additive effect on the increase in hepatic alkaline phosphatase activity, although the increase occurred earlier. These results suggest that: (a) cyclic AMP may act as an inducer of hepatic alkaline phosphatase: (b) the stimulation of hepatic alkaline phosphatase by cholera enterotoxin is mediated by cyclic AMP; (c) the rise in hepatic alkaline phosphatase following bile duct ligation is not mediated by cyclic AMP; (d) the same alkaline phosphatase in rat liver may be induced by two (or more) mechanisms, only one of which requires cyclic AMP.
Adenylyl Cyclases↗
Effects of cortisone administration on rat liver mitochondria. Support for the concept of mitochondrial fusion.
The present studies investigate the basis for the marked increase in mitochondrial size and approximately reciprocal decrease in mitochondrial number which have been observed in the livers of rats treated with cortisone acetate. Comparisons of the content and specific activity of mitochondrial DNA in the livers of control and cortisone-treated animals prelabeled with radioactive thymidine support the possibility that these changes in mitochondrial size and number are the result of a process of mitochondrial fusion. A consideration of various conditions now known to result in the formation of large mitochondria in other systems suggests that interference with mitochondrial respiration may provide a stimulus for such a process. The biochemical approach described in the present study may prove useful in investigating the origin of large mitochondria in other systems as well.
Animals↗
Differential sensitivity of nuclear and mitochondrial DNA synthesis to suppression by cortisone treatment.
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Aging↗
Stimulation of intestinal mucosal adenyl cyclase by cholera enterotoxin and prostaglandins.
The effects of several prostaglandins (PG) and a highly purified preparation of cholera enterotoxin (CT) on intestinal mucosal adenyl cyclase activity and the effect of CT on intestinal mucosal cyclic 3',5'-adenosine monophosphate concentration were determined in guinea pig and rabbit small intestine and were correlated with the effects of the same agents on ion transport. Adenyl cyclase activity, measured in a crude membrane fraction of the mucosa, was found at all levels of the small intestine with the highest activity per milligram protein in the duodenum. The prostaglandins, when added directly to the assay, increased adenyl cyclase activity; the greatest effect (2-fold increase) was obtained with PGE(1) (maximal effect at 0.03 mM) and PGE(2). The prostaglandins also increased short-circuit current (SCC) in isolated guinea pig ileal mucosa, with PGE(1) and PGE(2) again giving the greatest effects. The prior addition of theophylline (10 mM) reduced the subsequent SCC response to PGE(1) and vice versa. It was concluded, therefore, that the SCC response to PGE(1), like the response to theophylline, represented active Cl secretion. CT increased adenyl cyclase activity in guinea pig and rabbit ileal mucosa when preincubated with the mucosa from 1 to 2.5 hr in vitro or for 2.5 hr in vivo but not when added directly to the assay. The increments in activity caused by PGE(1) and NaF were the same in CT-treated and control mucosa. Cyclic 3',5'-AMP concentration in rabbit ileal mucosa was increased 3.5-fold after a 2 hr preincubation with CT in vitro. Phosphodiesterase activity in the crude membrane fraction of the mucosa was unaffected by either CT or PGE(1). A variety of other agents including insulin, glucagon, parathormone, thyroid-stimulating hormone, L-thyroxine, thyrocalcitonin, vasopressin, and epinephrine all failed to change adenyl cyclase activity. It is concluded that CT and certain prostaglandins produce small intestinal fluid secretion by increasing mucosal adenyl cyclase activity, thereby stimulating an active secretory process.
Adenine Nucleotides↗
Effect of cortisone treatment on the active transport of calcium by the small intestine.
It is generally recognized that glucocorticoid administration may diminish calcium absorption in vivo as well as the active transport of calcium by the intestine in vitro. Recent studies by others have emphasized the possibility of an alteration in the metabolism of vitamin D to 25-hydroxycholecalciferol in accounting for the steroid effects on calcium absorption. The results obtained in the present studies fail to support this hypothesis. The present studies confirm that the administration of cortisone or other glucocorticoids to the rat interferes with the active transport of calcium by duodenal gut sacs in vitro. This abnormality is not due to an alteration in the permeability of the intestine to calcium, and it cannot be corrected by the administration of either massive doses of vitamin D(2) or modest doses of 25-hydroxycholecalciferol. Experiments concerned with the effects of cortisone on the level of the vitamin D-dependent duodenal calcium-binding protein, the amount of bioassayable vitamin D activity in the mucosa, and the distribution and metabolism of (3)H-vitamin D(3), did not provide evidence in favor of a harmone-related defect in either the localization of vitamin D or its metabolism to 25-hydroxycholecalciferol. Alterations in the transport of iron and D-galactose, not dependent on vitamin D, suggest that cortisone treatment may be responsible for more than a simple antagonism to the effects of vitamin D. The results of the present studies indicate that cortisone administration affects the cellular mechanisms mediating calcium transport in a manner that is opposite to the effects of vitamin D, but seems to be independent of any direct interaction with the parent vitamin or its metabolites. If a disorder in vitamin D metabolism is at all involved, it is at a step subsequent to 25-hydroxylation.
Animals↗
Sedimentation properties of rat liver mitochondria. Effects of cortisone treatment.
A prediction of the velocity of sedimentation of rat liver mitochondria in sucrose gradients is made on the basis of recent measurements of the size of isolated mitochondria suspended in sucrose medium and the model proposed by Bentzel and Solomon to describe the osmotic behavior of mitochondria. The experimentally observed velocity is extremely close to the predicted value and confirms by a different approach the estimate of mitochondrial volume made by Baudhuin and Berthet on the basis of electron microscopic measurements. Because cortisone treatment of rats is known to result in a marked increase in mitochondrial size as observed under the electron microscope, mitochondria were co-isolated from livers of control and cortisone-treated animals, and the sedimentation behavior of the mixtures was examined by sucrose density gradient centrifugation. Mitochondria from cortisone-treated animals were found to sediment 1.4 times as rapidly as those from control animals, indicating that their increased size cannot entirely be due to an increased imbibition of fluid from the surrounding sucrose medium, and that the change in size must at least in part be due to a change in content of nondiffusible mitochondrial components. Although the increase in sedimentation velocity of mitochondria from cortisone-treated animals is striking, it is less than that predicted solely on the basis of their size relative to that of control mitochondria. It is concluded that the increases in mitochondrial size and content of nondiffusible components produced by cortisone treatment are accompanied by alterations in mitochondrial composition as well.
Animals↗
Effect of renal insufficiency on the active transport of calcium by the small intestine.
The intestinal absorption of calcium is often depressed in patients with chronic renal insufficiency. Furthermore, the malabsorption of calcium and the osteodystrophy which occur in association with chronic renal disease are often "resistant" to vitamin D; the basis for this resistance remains uncertain however. Recent studies by others have emphasized the role of an abnormality in the metabolism of vitamin D in accounting for the alterations in the calcium absorption and the apparent vitamin D-resistance which accompany the uremic syndrome. The present studies with an experimentally uremic animal model demonstrate a defect in the active transport of calcium by duodenal gut sacs in vitro. This abnormality is not due to the semistarvation associated with renal insufficiency and cannot be corrected by the administration of physiologic amounts of vitamin D(3): it is reversed by massive doses of the vitamin. Neither the metabolism of vitamin D(3) nor the levels of calcium binding protein activity in the duodenal mucosa are affected by renal insufficiency under the conditions employed in the present studies. The results of the present studies strongly suggest that in addition to the recently proposed mechanism involving an interference with the metabolism of vitamin D renal insufficiency also affects the cellular mechanisms for calcium transport in a manner which, while opposite in direction to that of vitamin D, is independent of a direct interaction with the vitamin or its metabolites.
Animals↗
Centrilobular hepatic necrosis and acute renal failure in "solvent sniffers".
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Acute Disease↗
The nature of vitamin D resistance in experimental uremia.
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Animals↗
Effects of vitamin D and steroid hormones on the active transport of calcium by the intestine.
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Adaptation, Physiological↗