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Biomedical subjects

M Donowitz

Publications and source records attributed to M Donowitz.

At least 127 records · Page 7Linked to original sources
PubMed1986

Suggestion of a role for calmodulin and phosphorylation in regulation of rabbit ileal electrolyte transport: effects of promethazine.

Suggestion of a role for protein phosphorylation in the regulation of intestinal active NaCl transport was found by studying the effects of low concentrations of promethazine on Ca2+-calmodulin (CaM)-dependent protein phosphorylation of ileal microvillus membranes and on active ileal electrolyte transport. Ca2+-CaM increased the phosphorylation of six microvillus peptides (Mr 137,000, 116,000, 77,000, 58,000, 53,000, and 50,000) in a concentration-dependent manner. Promethazine inhibited the Ca2+-CaM-induced increases in each of these phosphorylations. The effect of promethazine was concentration dependent, with concentrations of 5-12 microM (mean 8 microM) causing 50% inhibition. Promethazine also caused a concentration-dependent increase in net Cl absorption and decrease in the ileal short-circuit current, with 9 microM promethazine causing a change in short-circuit current 50% of maximum. The promethazine effect on microvillus membrane phosphorylation was specific, since neither cAMP- and cGMP-induced phosphorylation in the microvillus membrane nor the stimulation by Ca2+-CaM of myosin light chain kinase phosphorylation of myosin light chain were affected by promethazine. The similar, and unusual sensitivity to low concentrations of promethazine on ileal microvillus membrane phosphorylation increased by Ca2+-CaM and on ileal electrolyte transport is consistent with Ca2+-CaM-dependent microvillus membrane phosphorylation being involved in the regulation of active electrolyte transport in ileal absorptive cells.

Animals↗
PubMed1985

Secretory hormones of Entamoeba histolytica.

Watery diarrhoea as distinct from dysentery is occasionally seen in intestinal amoebiasis, suggesting a component of intestinal secretion. To study the pathogenesis of this watery diarrhoea, we evaluated the effect of lysates of Entamoeba histolytica on active intestinal electrolyte transport using rabbit ileum and rat colon studied by the Ussing chamber-voltage clamp technique. Amoebic lysates added to the ileal and colonic mucosal surfaces did not alter electrolyte transport; in contrast, addition to the ideal and colonic serosal surfaces caused an increase in short-circuit current which was transient in the ileum but more prolonged in the colon. This increase in current corresponded to inhibition of active Na+ and Cl- absorption and apparent stimulation of Cl- secretion in rat colon. In rabbit ileum, the short-circuit current response was: (1) dependent on serosal Ca2+, (2) inhibited by serosally applied verapamil, (3) associated with reversible desensitization, and (4) only partially inhibited by heating. These characteristics were similar to those of a series of neurohumoral substances present in mammalian intestinal mucosa which affect active electrolyte transport by increasing the permeability of the basolateral membrane to Ca2+. Substances with these properties identified include serotonin, substance P and neurotensin. All three substances were shown to be present in amoebic lysates by radioimmunoassay. Serotonin was also present by high performance liquid chromatography (HPLC) and thin layer chromatography, and neurotensin by HPLC. Prostaglandins were not present by radioimmunoassay. Attempts were made to determine which of these neurohumoral substances contributed to the changes in intestinal transport caused by amoebic lysates. Serotonin was thought to be involved, from the inhibition of the transport effect of amoebic lysate on rat colon by anti-serotonin antibody and by bufotenine, which inhibits the effect of serotonin on ion transport. Prostaglandins also appeared to be involved, since pretreatment with PGE2 or indomethacin inhibited the effects of amoebic lysate on transport. We conclude that amoebae contain neurohumoral substances, including serotonin, neurotensin and substance P, which may be important in the intestinal secretion caused by amoebae. Serotonin appears partially responsible for the intestinal secretion. In addition, amoebae may induce prostaglandin synthesis by the intestinal mucosa which could also contribute to the secretory response. The relation between the neurohumoral substances which act by Ca2+ and the prostaglandins presumably caused to be synthesized in the intestinal mucosa is not known.

Animals↗
PubMed1985

Ca2+ channel blockers stimulate ileal and colonic water absorption.

The effects of calcium channel blockers on water transport in the rat ileum and distal colon were studied in vivo using the single-pass perfusion technique. Parenteral but not intraluminal verapamil, and parenteral nifedipine increased ileal water absorption, with effects lasting at least 60 min. In contrast, i.p. verapamil had no effect on rat distal colonic water absorption, whereas intraluminal verapamil significantly stimulated colonic water absorption. Similarly, perfusing the rat descending colon with low-Ca2+ Ringer's-HCO3 stimulated colonic water absorption. Verapamil was not antisecretory because the theophylline-induced decrease in ileal water transport was similar in control animals and in animals pretreated with i.p. verapamil. In addition, nifedipine stimulated active Na and Cl absorption in rabbit ileum. These studies demonstrate that the Ca2+ channel blockers verapamil and nifedipine stimulate basal absorption of water in rat ileum and distal colon in vivo, and stimulate active Na and Cl absorption in rabbit ileum in vitro. The verapamil stimulation of colonic water absorption from the luminal surface was duplicated by perfusion with a low-Ca2+ bathing solution. This suggests the presence of apical membrane Ca2+ channels in rat colon, which appear to be involved in regulation of basal water transport, and that these Ca2+ channels are in a partially open state under basal conditions. Because verapamil stimulates absorption systemically (ileum) as well as intraluminally (colon), Ca2+ channel blockers have properties that might be useful in treatment of diarrheal diseases.

Animals↗
PubMed1985

Extrahepatic portal venous thrombosis: frequent recognition of associated diseases.

Twelve patients with obstruction of the extrahepatic portal venous system were seen at the Tufts-New England Medical Center between 1970 and 1979; a cause for the portal vein thrombosis was detected in 11. These included pancreatic disease (4); hematologic disorders (2); postoperative complications of laparotomy (3); transhepatic gelfoam embolization of the portal vein (1); and exchange transfusion via the umbilical vein (1). Clinical features included frequent self-limited episodes of bleeding from esophageal or gastric varices; and no characteristic or clinically helpful laboratory findings. The diagnosis was usually made in patients by identifying clots in the portal vein on selective angiography of the celiac and/or superior mesenteric arteries in which the venous phase was examined. Attempts at surgical correction were largely unsuccessful. Further thrombotic episodes occurred in three patients, and led to death in one. Two patients were given chronic anticoagulation with Coumadin and Persantin for 1 and 11/2 years, respectively without further thrombosis or gastrointestinal bleeding. However, it not yet possible to assess the risks and benefits of such therapy.

Adolescent↗
PubMed1985

Studies on role of calmodulin in Ca2+ regulation of rabbit ileal Na and Cl transport.

To study the role of calmodulin in regulation of rabbit ileal active electrolyte transport by Ca2+, the effects of the Ca2+-calmodulin antagonist naphthalenesulfonamides W12 and W13 and the weak Ca2+-calmodulin antagonist promethazine, a phenothiazine, were studied on basal ileal Na and Cl transport and on secretion stimulated by Ca2+ and by cAMP. The naphthalenesulfonamides and promethazine all stimulated basal ileal Na and Cl absorption. The stimulation of Na absorption was dependent on Cl in the bathing solutions, and the stimulation of Cl absorption was Na dependent. This suggests that the transport process stimulated was the neutral, linked NaCl absorptive process. W13, which is a better Ca2+-calmodulin antagonist but has similar hydrophobic properties to W12, stimulated active ileal absorption at a lower concentration than W12, suggesting that the naphthalenesulfonamide-induced stimulation of active ileal absorption was not due to the hydrophobic properties of these drugs but could be due to their effects on the calcium-binding protein calmodulin. W12, W13, and promethazine did not alter paracellular transport, not affecting dilution potentials or structural features of the paracellular pathway. W12 and W13 did not decrease the changes in active ileal Na and Cl transport caused by increasing ileal cAMP content or intracellular Ca2+. This suggests that calmodulin is not directly involved in the active electrolyte secretion caused by increased intestinal Ca2+ or cAMP.

Animals↗
PubMed1985

Role of calcium in cholinergic stimulation of lacrimal gland protein secretion.

To characterize the role of Ca2+ in cholinergic stimulation of lacrimal gland protein secretion, the effects of inhibitors of cellular Ca2+ handling on protein secretion were investigated. Protein secretion was measured from rat exorbital glands using either pieces of gland in perifusion or acini isolated by collagenase digestion. Peroxidase was used as a measure of protein secretion. An inhibitor of Ca2+ influx via voltage sensitive Ca2+ channels (verapamil) at 10(-5) and 5 X 10(-5) M did not alter protein secretion stimulated by the cholinergic agonist carbachol at 10(-5) M. Inhibition of Ca2+ efflux via Na+/Ca2+ exchange by removal of extracellular Na+ or by inhibition of Na+-K+-ATPase activity using ouabain (10(-3) M) or extracellular K+ removal did not stimulate protein secretion. In contrast, inhibition of Ca2+ release from intracellular stores with TMB-8 at 100 micron completely blocked protein secretion stimulated by carbachol at 10(-5) M. Similarly, the Ca2+/calmodulin (CaM) antagonists W-13 and W-12 decreased carbachol-induced protein secretion with potencies similar to those which inhibit Ca2+/CaM dependent processes. We conclude that cholinergic agonists stimulate lacrimal gland protein secretion primarily by mobilizing Ca2+ from intracellular stores and that one mechanism by which this Ca2+ could activate secretion is in conjunction with calmodulin.

Animals↗
PubMed1984

Ca2+-calmodulin-, cyclic AMP- and cyclic GMP-induced phosphorylation of proteins in purified microvillus membranes of rabbit ileum.

Evidence is available to suggest that Ca2+-calmodulin and cyclic nucleotides are involved in the regulation of ion transport in rabbit ileum. Since both Ca2+-calmodulin and cyclic nucleotides exert many of their effects by phosphorylation, the effects of Ca2+-calmodulin and cyclic nucleotides on phosphorylation of purified microvillus membrane from rabbit ileal mucosa were evaluated. Ca2+-calmodulin increased phosphorylation of five microvillus-membrane peptides, with Mr values of 137000, 77000, 58000, 53000 and 50000. The increases in phosphorylation caused by Ca2+-calmodulin were: Mr-137000 peptide, 111 +/- 26%; Mr-77000 peptide, 71 +/- 17%; Mr-58000 peptide, 51 +/- 8%; Mr-53000 peptide, 113 +/- 20%. These increases were maximal at 1 microM-calmodulin and 0.3-0.9 microM free Ca2+; concentrations of Ca2+ causing half-maximal effects on phosphorylation for the different peptides were 0.06-0.12 microM. Cyclic AMP and cyclic GMP increased phosphorylation of two peptides, of Mr 137000 and 85000. The concentrations of cyclic nucleotides giving half-maximal phosphorylation of the Mr-137000 peptide were 0.3 microM-cyclic AMP and 4.6 microM-cyclic GMP, and for the Mr-85000 peptide, 3.9 microM-cyclic AMP and 0.05 microM-cyclic GMP. The maximal increase in phosphorylation of the Mr-137000 peptide was 200% for cyclic AMP and 95% for cyclic GMP, and that of the Mr-85000 peptide was 220% for cyclic AMP and 120% for cyclic GMP. These studies demonstrate the existence of Ca2+-calmodulin-, cyclic AMP- and cyclic GMP-dependent protein kinases and substrate proteins in purified rabbit ileal microvillus membranes and that Ca2+ can regulate phosphorylation of these proteins over the presumed physiological concentration range of cytosol free Ca2+.

Animals↗
PubMed1984

Acid and isoproterenol cause serotonin release by acting on opposite surfaces of duodenal mucosa.

Although most bodily serotonin (5-HT) is stored in intestinal enterochromaffin (EC) cells, the mechanism of its release is only now being elucidated. It has previously been reported that such stimuli as luminal acidification or exposure of both sides of a rabbit duodenal mucosal sheet to certain autonomic agonists stimulate release from the mucosal surface in the Ussing chamber model. The hypothesis in the present study is that acid acts only on the mucosal surface, whereas neural receptor agonists and antagonists act specifically on the serosal surface, as would be predicted by the location of acid in the gut lumen and nerve terminals at the bases of the EC cells. 5-HT release was measured by radioimmunoassay from the mucosal surface bathing solution. Duodenal mucosal sheets were exposed separately on the mucosal or serosal surfaces to acid (citric phosphate buffer, pH 5) or to isoproterenol (10(-5) M). The effect of atropine (10(-6) M) and propranolol (10(-6) M) on acid-stimulated mucosal release was studied by combining luminal acid stimulation with one of these antagonists, on either the mucosal or serosal surface. The results demonstrate significant (P less than 0.01) mucosal serotonin release (56 +/- 9 ng cm-2 hr-1) only with mucosal acidification. On the other hand, isoproterenol causes significant (P less than 0.05) serotonin release (12.4 +/- 3 ng cm-2 hr-1) only when introduced onto the serosal surface. Finally, the antagonists, atropine and propranolol, blocked acid-stimulated serotonin release only when added to the serosal surface. Since acid-induced serotonin release has been shown to be partially mediated by cholinergic and beta-adrenergic mechanisms, these findings suggest interconnection of mucosal acid receptors with submucosal neurons which mediate serotonin release by acting on the basal surface of the enterochromaffin cell.

Animals↗
PubMed1984

Cyclic nucleotide-dependent enzyme secretion in the rat lacrimal gland.

To characterize the role of cyclic nucleotides in secretion of enzymes by the lacrimal gland, pieces of rat exorbital glands were perfused with (1) 8-bromoadenosine-3',5'-cyclic monophosphate (8 Br cyclic AMP), (2) 8-bromoguanosine-3',5'-cyclic monophosphate (8 Br cyclic GMP), (3) forskolin, a stimulator of adenylate cyclase activity, (4) 3-isobutyl-1-methylxanthine (IBMX), an inhibitor of phosphodiesterase activity, or (5) carbachol, a cholinergic agonist. As a measure of enzyme secretion, timed collections of the perifusate effluent were analysed for peroxidase, an enzyme secreted by the lacrimal gland. Control peroxidase secretion was 0.3-0.9 (u./min per milligram protein). Peroxidase secretion was stimulated by 8 Br cyclic AMP (1 mM), but not by 8 Br cyclic GMP (1 mM). A 2-fold increase was detected. Peroxidase secretion was also stimulated by forskolin (60 microM), IBMX (1 mM), and the cholinergic agonist carbachol, which all stimulated peroxidase secretion 2-or 3-fold. The effect of maximally effective concentrations of IBMX (1 mM) and carbachol (0.1 mM) on secretion was additive. Finally, Ca2+ depletion in the presence of EGTA (1 mM) inhibited both IBMX-and carbachol-induced secretion by 45% and 60% respectively. We conclude that cyclic AMP, but not cyclic GMP, can stimulate lacrimal gland enzyme secretion. Cyclic AMP appears to utilize a pathway separate from but convergent with cholinergic agonists.

1-Methyl-3-isobutylxanthine↗
PubMed1984

Effect of Senokot on rat intestinal electrolyte transport. Evidence of Ca++ dependence.

The mechanism of action of Senokot, a widely used laxative, has not been established. Senokot was given orally to rats 8-14 h before intestinal water and electrolyte transport were studied. Senokot significantly decreased colonic and jejunal water absorption measured in vivo by the single-pass perfusion technique. The Senokot changes were not associated with changes in jejunal or colonic histology or adenylate cyclase activity or colonic cyclic adenosine monophosphate content. Senokot also altered active electrolyte transport in rat descending colon as measured by the Ussing chamber-voltage clamp technique. These changes consisted of an increase in short-circuit current and a decrease in active Na and Cl transport that was due to a decrease in the mucosal-to-serosal fluxes. The changes in active electrolyte transport were dependent on Ca++ in the serosal but not the mucosal bathing solution. In contrast, addition of 10(-4) M verapamil to the serosal surface did not alter the Senokot effect. In spite of a dependence on serosal Ca++, Senokot did not alter 45Ca++ entry across the colonic serosal surface. The phospholipase A2 inhibitor quinacrine (10(-4) M) also did not alter the effect of Senokot on colonic Na and Cl transport. Senokot alters active colonic Na and Cl transport via a presently unknown mechanism that is dependent on serosal Ca++.

Adenylyl Cyclases↗
PubMed1984

Enteric neural pathways inhibitory to rabbit duodenal serotonin release.

Previous studies have shown that adrenergic and cholinergic neural pathways mediate duodenal serotonin (5-HT) release from the mucosal surface after a luminal acid stimulus. To examine the overall neural contribution to 5-HT release, we studied the effects of tetrodotoxin (TTX), a nerve-conduction blocker, added to mucosal and/or serosal surfaces of rabbit duodenal mucosa in a modified Ussing chamber at both neutral and acid (pH 5) luminal pH. A specific radioimmunoassay was used to measure 5-HT. TTX (2 X 10(-7) M) increased mucosal 5-HT release significantly at luminal pH 7.4 and 5 when it was added to both mucosal and serosal surfaces (2.7- and 1.9-fold at pH 7.4 and pH 5, respectively; p less than 0.05 for each) or when it was added to the serosal surface only (3.4- and 1.8-fold; p less than 0.02 and p less than 0.01, respectively). TTX added only to the mucosal surface, however, reduced 5-HT release by 48.5% at pH 5 (p less than 0.01) and had no effect at pH 7.4. Since adenosine triphosphate (ATP) and vasoactive intestinal polypeptide are the two proposed mediators of a nonadrenergic, noncholinergic inhibitory neural system, we studied the effects of each on mucosal 5-HT release. While vasoactive intestinal polypeptide had no significant effect at either pH, ATP (10(-7)M) reduced acid-stimulated 5-HT release by approximately 50% (p less than 0.02). We conclude that a nonadrenergic, noncholinergic neural pathway that is inhibitory to mucosal 5-HT release resides primarily on the basal surface of mucosal cells. ATP is the most likely neurotransmitter involved in this response.

Adenosine Triphosphate↗
PubMed1983

Entamoeba histolytica causes intestinal secretion: role of serotonin.

Lysates of the protozoan parasite Entamoeba histolytica altered active electrolyte transport when present on the serosal surface of rabbit ileum and rat colon. The lysate-induced effects on electrolyte transport were similar to those caused by serotonin, and were blocked by bufotenine, an analog known to inhibit the action of serotonin. The transport effects were partially inhibited by antibody to serotonin. The amebic lysates were shown to contain serotonin by radioimmunoassay, high-performance liquid chromatography, and thin-layer chromatography. These results suggest that the serotonin present in Entamoeba histolytica may be important in the diarrhea seen in amebiasis.

Amebiasis↗
PubMed1983

Ca2+ in the control of active intestinal Na and Cl transport: involvement in neurohumoral action.

Intracellular Ca2+ is a regulator of active intestinal Na and Cl transport. Most studies have been done with rabbit ileum. Increasing intracellular Ca2+ decreases active Na and Cl absorption and/or stimulates active Cl secretion; lowering intracellular Ca2+ stimulates Na and Cl absorption. Based on studies with microvillus membrane vesicles from rabbit ileum, a direct effect of Ca2+ and calmodulin on linked Na and Cl uptake is established. Intracellular Ca2+ and cAMP affect the same transport processes and act in a nonadditive manner. Intracellular Ca2+ does not act by changing intestinal cAMP or cGMP contents, and increasing cAMP mobilizes intracellular Ca2+. Whether this Ca2+ is involved in regulation of ion transport is not known. The aspects of Ca2+ handling identified as involved in regulation of active intestinal Na and Cl transport include entry of Ca2+ across the basolateral membrane, mobilization of Ca2+ from intracellular stores, and involvement of the Ca2+-binding protein calmodulin. Several neurohumoral substances alter intestinal transport by Ca2+-dependent mechanisms and appear to act primarily by increasing (serotonin, carbachol, substance P, and neurotensin) or decreasing (dopamine) Ca2+ entry across the basolateral membrane of intestinal epithelial cells.

Animals↗
PubMed1983

Dantrolene and basal ileal sodium and chloride transport: involvement of calcium stores.

The effect of dantrolene on active ion transport in rabbit ileum was determined using the Ussing chamber short-circuiting technique. Dantrolene prevents the release of calcium from intracellular stores in skeletal muscle and was used to probe the role of intracellular calcium stores in intestinal ion transport. A saturated solution of dantrolene (approx 25 microM) decreased ileal short-circuit current and potential difference, increased conductance and mucosal-to-serosal and net Na and Cl fluxes, but did not alter serosal-to-mucosal Na and Cl fluxes. The dantrolene stimulation of active Na and Cl absorption was specific since it did not alter glucose-dependent Na absorption, transport changes caused by Ca2+ ionophore A23187, or the increase in short-circuit current caused by dibutyryl cAMP or theophylline. These effects were associated with an increase in total ileal calcium content and a decreased rate of 45Ca2+ efflux without any change in 45Ca2+ influx from the serosal or mucosal surfaces. These findings are consistent with an effect of dantrolene to stimulate active ileal Na and Cl absorption by a mechanism involving lowered cytosol Ca2+ levels and compatible with trapping calcium in intracellular stores. It thus appears as if intracellular calcium stores have an important role in the control of basal ion transport in the intestine.

Animals↗