Preoperative cow-side lactatemia measurement predicts negative outcome in Holstein dairy cattle with right abomasal disorders.

The objectives of the current study were (1) to determine the gain in prognostic accuracy of preoperative l-lactate concentration (LAC) measured on farm on cows with right displaced abomasum (RDA) or abomasal volvulus (AV) for predicting negative outcome; and (2) to suggest clinically relevant thresholds for such use. A cohort of 102 cows with on-farm surgical diagnostic of RDA or AV was obtained from June 2009 through December 2011. Blood was drawn from coccygeal vessels before surgery and plasma LAC was immediately measured by using a portable clinical analyzer. Dairy producers were interviewed by phone 30 d following surgery and the outcome was determined: a positive outcome if the owner was satisfied of the overall evolution 30 d postoperatively, and a negative outcome if the cow was culled, died, or if the owner reported being unsatisfied 30 d postoperatively. The area under the curve of the receiver operating characteristic curve for LAC was 0.92 and was significantly greater than the area under the curve of the receiver operating characteristic curve of heart rate (HR; 0.77), indicating that LAC, in general, performed better than HR to predict a negative outcome. Furthermore, the ability to predict a negative outcome was significantly improved when LAC measurement was considered in addition to the already available HR data (area under the curve: 0.93 and 95% confidence interval: 0.87, 0.99). Important inflection points of the misclassification cost term function were noted at thresholds of 2 and 6 mmol/L, suggesting the potential utility of these cut-points. The 2 and 6 mmol/L thresholds had a sensitivity, specificity, positive predictive value, and negative predictive value for predicting a negative outcome of 76.2, 82.7, 53.3, and 93.1%, and of 28.6, 97.5, 75, and 84%, respectively. In terms of clinical interpretation, LAC ≤2 mmol/L appeared to be a good indicator of positive outcome and could be used to support a surgical treatment decision. The treatment decision for cows with LAC between 2 and 6 mmol/L, however, would depend on the economic context and the owner's attitude to risk in regard to potential return on its investment. Finally, performing a surgical correction on commercial cows with RDA or AV and a LAC ≥6 mmol/L appeared to be unjustified and these animals should be culled based on their high probability of negative outcome.

Ulcerative colitis and adenocarcinoma of the colon in G alpha i2-deficient mice.

G proteins are involved in cellular signalling and regulate a variety of biological processes including differentiation and development. We have generated mice deficient for the G protein subunit alpha i2 (G alpha i2) by homologous recombination in embryonic stem cells. G alpha i2-deficient mice display growth retardation and develop a lethal diffuse colitis with clinical and histopathological features closely resembling ulcerative colitis in humans, including the development of adenocarcinoma of the colon. Prior to clinical symptoms, the mice show profound alterations in thymocyte maturation and function. The study of these animals should provide important insights into the pathogenesis of ulcerative colitis as well as carcinogenesis.

Preoperative and postoperative L-lactatemia assessment for the prognosis of right abomasal disorders in dairy cattle.

BACKGROUND: Preoperative L-lactatemia and heart rate have been suggested as prognostic indicators of outcome for cows with right dilatation of the abomasum or volvulus (RDA/AV). However, postoperative L-lactatemia has not been assessed as a potential prognostic tool. OBJECTIVES: To determine the prognostic value of postoperative L-lactatemia (LAC2 ), duration of treatment (Dt), relative L-lactatemia difference (compared with preoperative L-lactatemia [LAC1 ]) ([LAC2 - LAC1 ]/LAC2 ) and change in L-lactate over time ([LAC2 - LAC1 ]/Dt) as compared to preoperative findings (LAC1 and heart rate [HR]) as prognostic factors in dairy cows with RDA/AV. ANIMALS: A total of 41 dairy cows were included: 19 with AV and 22 with RDA; 11 cows had a negative outcome (NO) and 30 cows had a positive outcome (PO) based on telephone follow-up with owners 30 days after surgery. METHODS: Prospective cohort study. Analysis was performed using logistic regression and comparison of area under the receiver operating characteristics curve (AUC) using nonparametric tests. RESULTS: LAC1 > 1.4 mmol/L or LAC2 > 2.2 mmol/L had the same accuracy with sensitivity of 100% (95% CI, 75.1-100%) and specificity of 80% (95% CI, 61.4-92.3%) for predicting NO. The relative L-lactatemia difference ([LAC2 - LAC1 ]/LAC1 ) or lactate kinetics ([LAC2 - LAC1 ]/Dt) were not associated with prognosis. The AUC of the preoperative model (which included HR and lnLAC1 ) was 0.92 (95% CI, 0.83-1.0) and that of the postoperative model (including only lnLAC2 ) was 0.95 (95% CI, 0.88-1.0); these were not significantly different. CONCLUSIONS AND CLINICAL IMPORTANCE: Postoperative L-lactatemia is helpful to predict outcome in cows with RDA/AV. The short-term change in blood L-lactate is not a useful prognostic indicator, at least during the period of time spent on the farm for surgery and treatment.

Ca(2+)-calmodulin regulates receptor-operated Ca(2+) entry activity of TRPC6 in HEK-293 cells.

Mammalian homologues of the Drosophila transient receptor potential channel (TRPC) are involved in Ca(2+) entry following agonist stimulation of nonexcitable cells. Seven mammalian TRPCs have been cloned but their mechanisms of activation and/or regulation are still the subject of intense research efforts. It has already been shown that calmodulin (CaM) can regulate the activity of Drosophila TRP and TRPL and, more recently, CaM has been shown to interact with mammalian TRPCs. In this study, TRPC6 stably transfected into HEK-293 cells was used to investigate the possible influence of CaM on TRPC6-dependent Ca(2+) entry. Overexpression of TRPC6 in mammalian cells is known to enhance agonist-induced Ca(2+) entry, but not thapsigargin-induced Ca(2+) entry. Here, we show that CaM inhibitors (calmidazolium and trifluoperazine) abolish receptor-operated Ca(2+) entry (ROCE) without affecting thapsigargin-operated Ca(2+) entry and that the activity of CaM is dependent on complexation with Ca(2+). We also show that Ca(2+)-CaM binds to TRPC6 and that the binding can be abolished by CaM inhibitors. These results indicate that CaM is involved in the modulation of ROCE.

Two Ca2+ transport systems are distinguished on the basis of their Mg2+ dependency in a post-nuclear particulate fraction of bovine adrenal cortex.

Inositol 1,4,5-trisphosphate (InsP3) is a second messenger responsible for Ca2+ release from an internal store whose nature and location remains undefined. To get more information on this intracellular Ca2+ store, a post-nuclear particulate fraction was prepared from bovine adrenal cortex and its Ca2+ uptake and release activities were monitored with the fluorescent indicator Fura-2. In the presence of Mg2+ (2 mM), the particulate preparation showed high ATP-dependent Ca2+ sequestering activity and decreased the ambient Ca2+ concentration to about 150 nM. In the absence of Mg2+, Ca2+ was still sequestered but less efficiently, reaching a level around 170 nM. In the presence of Mg2+, the Ca2+ released by a maximal dose of InsP3 (2 microM) was completely resequestered whereas in the absence of Mg2+, no resequestration occurred even after complete degradation of InsP3. The use of selective agents such as oligomycin, saponin, ionomycin and biliary salts indicated that Ca2+ was stored in three different pools which are distinct from the mitochondria and from inside-out membrane vesicles. Our data also indicate that InsP3 releases Ca2+ from a pool which is filled up by a Mg2(+) -dependent Ca2+ ATPase.

The important discrepancy between the apparent affinity observed in Ca2+ mobilization studies and the Kd measured in binding studies is a consequence of the quantal process by which inositol 1,4,5-trisphosphate releases Ca2+ from bovine adrenal cortex microsomes.

Inositol 1,4,5-trisphosphate (InsP3) is a second messenger responsible for Ca2+ release from a non-mitochondrial intracellular store. An important discrepancy has been observed between the affinity measured in binding studies (Kd) and the apparent affinity obtained in Ca2+ mobilization studies (EC50). It has been proposed that this discrepancy could be due to different experimental conditions used for Ca2+ mobilization studies and for InsP3 binding studies. With the fluorescent indicator Fura-2, we studied InsP3-induced Ca2+ release activity at 7 degrees C and at 37 degrees C, in bovine adrenal cortex microsomes. Under both conditions, the Ca2+ releasing effect of InsP3 (1 microM) was completed within about 2 s, as a result of the quantal process of InsP3 receptor action. The apparent affinity (EC50) observed for InsP3-induced Ca2+ release at 7 degrees C and at 37 degrees C were 0.64 +/- 0.2 microM and 0.9 +/- 0.2 microM respectively. InsP3 degradation studies, at 37 degrees C, indicated that less than 10% of [3H]-InsP3 was degraded within the first 10 s of incubation. InsP3 association rates were evaluated, at low temperature, with increasing concentrations of [3H]-InsP3. These kinetic studies revealed a direct relationship between the initial rate of association (Vi) and InsP3 concentration. From this relationship, we evaluated that the concentration of InsP3 needed to occupy half of the binding sites within the first second of incubation was 271 nM. We conclude that the discrepancy between Kd and EC50 is related to a kinetic constraint dictated by the quantal process by which InsP3 releases Ca2+.

Short-term desensitization of the angiotensin II receptor of bovine adrenal glomerulosa cells corresponds to a shift from a high to a low affinity state.

Angiotensin II (Ang II) is an important regulator of aldosterone production by bovine adrenal glomerulosa (BAG) cells. Ang II interacts with a specific receptor coupled to a guanyl nucleotide-binding protein (G protein) that controls the activity of phospholipase C. A primary culture of BAG cells was used to study short-term desensitization of the Ang II receptor. After short exposures to Ang II, BAG cells lost some [125I]Ang II binding capacity. This loss was dependent on the duration of the pretreatment and on the concentration of Ang II used. A maximal loss of [125I]Ang II binding of 55 +/- 10% was observed after a pretreatment of 30 min with 30 nM Ang II. The EC50 was 1.3 +/- 0.6 nM (mean +/- SD of three experiments). The desensitization was readily reversible, since most of the binding capacity (higher than 90%) was recovered after a 60-min incubation, at 37 C, in the absence of Ang II. Scatchard studies revealed that the Ang II receptor of BAG cells exists under two affinity states with one dissociation constant of 0.2 nM and another dissociation constant of 1.5 nM. After a 30-min exposure of BAG cells to 10 nM Ang II, an important decrease of high affinity binding sites was observed. The maximal amount of binding sites was similar on control and desensitized cells (around 52,000 receptors per cell). GTP gamma S, a potent activator of G proteins, decreased [125I]Ang II binding to permeabilized BAG cells. This GTP gamma S effect was not observed on permeabilized BAG cells that had previously been desensitized with 10 nM Ang II. These results suggested that, similarly to GTP gamma S, short exposure to 10 nM Ang II caused the uncoupling of Ang II receptor from its G protein. DuP-753 (a selective AT1 angiotensin II type 1 receptor antagonist) markedly unhibited, whereas PD-123319 (a selective AT2 angioten II type 2 receptor antagonist) had no effect on Ang II receptor desensitization, indicating that the AT1 receptor subtype was responsible for the observed phenomenon. Pretreatment of BAG cells with staurosporine (a protein kinase C inhibitor) and R24571 (a calmodulin inhibitor) did not modify Ang II-induced desensitization of AT1 receptor.(ABSTRACT TRUNCATED AT 400 WORDS)

A 31P nuclear magnetic resonance in vivo study of cerebral ischaemia in the gerbil.

We have used the noninvasive method of 31phosphorus nuclear magnetic resonance (31P NMR) in vivo to follow changes in phosphorous metabolite concentrations and the intracellular pH in the right and left hemispheres and in the cerebellum of gerbil brains after the occlusion of the right carotid artery. Spatial resolution over the brain was possible using surface coils. Ligation, which is know to cause ischaemia in this species in the ipsilateral hemisphere, resulted in the diminution of phosphocreatine and adenine nucleotides and a decrease in tissue pH. Less acidification occurred in the contralateral hemisphere and in the cerebellum. The high-energy metabolite concentrations, phosphocreatine and adenosine triphosphate (ATP), declined in unison in the ischaemic region, in marked contrast to the sequence of events in skeletal muscle, in which phosphocreatine buffers against an immediate fall in ATP concentration. In a separate series of gerbils, 31P NMR spectra were followed for exactly 1 h after carotid ligation. The animals were then sacrificed and brain grey matter specific gravity was rapidly measured to assess the development of oedema. There was a clear correlation between abnormality of spectra and the presence of oedema. It cannot, however, be confidently asserted that a normal spectrum is never seen in oedematous gerbil brains. 31P NMR spectra specific gravity and histological changes shown by light microscopy have been correlated and show that useful signals are received from a depth of at least 4 mm or more from the 10-mm diameter coil.

Ionic currents underlying HTRP3 mediated agonist-dependent Ca2+ influx in stably transfected HEK293 cells.

hTrp3 is a human homologue of the Drosophila gene responsible for a transient receptor potential (trp) mutation. When stably expressed in HEK293 cells, hTrp3 formed ion channels that were active under resting conditions but could be further stimulated by carbachol or ATP via endogenous muscarinic or purinergic receptors, respectively. Agonist evoked currents reversed polarity near 0 mV in physiological ionic conditions and were associated with a significant increase in the current variance. These results suggest the involvement of a non-selective cation channel with relatively large unitary amplitude. Consistent with this, resolved unitary events had a conductance of approximately 60 pS in the negative voltage range and an extrapolated reversal potential near 0 mV.

Cerebral blood flow in dementia.

Twenty-four patients of comparable age, blood pressure, and degree of dementia were classified by an "Ischemic Score" based on clinical features into "multi-infarct" and "primary degenerative" dementia. Regional cerebral blood flow (CBF) was measured by the intracarotid xenon 133 method. Both groups showed a decreased proportion of rapidly clearing brain tissue (largely gray matter). Cerebral blood flow per 100 gm brain per minute was normal in the primary degenerative group but low in the multi-infarct group. This suggests the blood flow is adequate for metabolic needs of the brain in patients with primary degenerative dementia but inadequate for those with multi-infarct dementia. There was no correlation between degree of dementia and CBF in the primary degenerative group but an inverse relationship existed in the multi-infarct group. Reactivity of blood vessels to reduction of arterial carbon dioxide pressure was normal in both groups.

Comparing Evans' index and computerized axial tomography in assessing relationship of ventricular size to brain size.

Using pneumoencephalography and computerized axial tomography (EMI scanning) Polaroid pictures, the relationship between ventricular size and cerebral size was investigated in 35 patients. Evans' index was used for pneumoencephalograms, and planimetric measurement of the ventricular and cerebral cross-sectional areas was used for EMI scanning. The percentage ratio for the latter technique is termed VBR. The correlation coefficient between the two methods was 0.9510 (p less than 0.001).

Cerebrovascular carbon dioxide reactivity and conductance in patients awake and under general anesthesia.

Paired cerebral blood flow measurements at two levels of arterial carbon dioxide (CO2) tension were made in 36 awake patients and 85 patients under general anesthesia to measure the percent change in flow per mm Hg change in PaCO2 (CO2 reactivity). CO2 reactivity was significantly greater in the generally anesthetized (5.96) than in the awake group (3.65). In both groups, a linear correlation was found between mean percent CO2 reactivity and conductance (the reciprocal of resistance), up to a level of conductance of 0.85 in awake and 0.95 in unconscious patients. When the CO2 reactivity of 18 awake patients and 59 unconscious patients was compared with that of respective control groups, only 11 patients were found to have abnormal CO2 reactivities for their conductance levels.

Frequent involvement of the optic radiation in patients with acute isolated optic neuritis.

Magnetic resonance imaging revealed asymptomatic lesions in white matter regions corresponding with the optic radiations in 20 of 28 patients (71%) with clinically isolated optic neuritis. In contrast to the findings with symptomatic lesions, there was no relationship between the latency of the visual evoked potential and the presence of these asymptomatic posterior visual pathway lesions.

Focal signal hyperintensities in schizophrenia.

The presence of focal signal hyperintensities on MRI images of the brain was compared in 48 schizophrenic patients, 26 patients with bipolar disorder and 34 healthy controls. Significantly larger areas of brain were affected by focal signal hyperintensities, particularly in the frontal lobes, in the schizophrenic group compared to the bipolar group and the controls. Although the bipolar group had more such foci than controls, this difference did not reach statistical significance.

Sar1-p-benzoylphenylalanine-angiotensin, a new photoaffinity probe for selective labeling of the type 2 angiotensin receptor.

Previous photoaffinity labeling of angiotensin II (Ang) receptors with azidophenylalanine containing Ang analogs produced high yield labeling of a 60 kDa protein on bovine adrenocortical membranes. This preparation is mostly enriched in the type 1 Ang receptor (AT1) and AT1 selective ligands (L158,809) totally prevented labeling, therefore confirming the AT1 nature of the labeled protein. Our attempt to photolabel the type 2 Ang receptor (AT2) of human myometrium with [Sar1,D-Phe(N3)8]Ang was unsuccessful, revealing a high degree of photolabeling selectivity. An Ang analog, [Sar1,Bpa8]Ang (or BpaAng) was prepared containing the photosensitive amino acid p-benzoylphenylalanine (Bpa). This compound was a specific but non-competitive Ang antagonist on rabbit aorta with a pA2 of 8.5. It displayed good binding affinities for bovine adrenocortical membranes (Kd = 6.5 nM), a predominantly AT1 preparation, and for human myometrium membranes (Kd = 0.39 nM), a predominantly AT2 preparation. Photolabeling experiments with iodinated BpaAng showed that AT1 was not covalently labeled whereas AT2 was covalently labeled with high yield. Labeling specificity was verified with the AT2-selective ligand PD123319 and with the AT1-selective antagonist L158,809. Our results indicate that 125I-BpaAng is exclusively labeling AT2 sites. This compound should be a useful tool for further biochemical characterization of the AT2 binding site.

Patterns of disease activity in multiple sclerosis patients: a study with quantitative gadolinium-enhanced brain MRI and cytokine measurement in different clinical subgroups.

In this study we assessed the subclinical disease activity in 45 patients with primary progressive, secondary progressive or relapsing-remitting multiple sclerosis (MS). The patients had gadolinium-enhanced brain MRI scans, which were analysed using a semiquantitative method both for lesion load and for degree of enhancement. At the same time cerebrospinal fluid (CSF) and serum samples were collected and, from these, cytokine levels were measured in most cases by enzyme-linked immunoassay using commercially available kits. Enhancing lesions on MRI were found in 73% of the patients. The sensitivity of this test was greatly increased by our method of analysis as far as the primary progressive patients are concerned (70% vs 40% for conventional evaluation). CSF interleukin-1 beta (IL-1 beta) levels were above the normal range in 22% and IL-6 levels in 13% of patients, while tumour necrosis factor alpha (TNF-alpha) was undetectable or below the upper normal limits in all the samples tested. Serum IL-1 beta was above the normal limits in 40%, IL-6 in 42% and TNF-alpha in 7% of patients. No significant differences in cytokine profiles were found between the clinical subgroups. This study confirms the high sensitivity of gadolinium-enhanced MRI in detecting MS activity, which was further increased by our method of analysis. Longitudinal studies performed with more sensitive immunological techniques are needed to define better the relationship between cytokine, clinical and MRI data in MS patients.

Implication of phospholipase C in the steroidogenic action of angiotensin II.

Angiotensin II (AII) is a major regulator of aldosterone synthesis and secretion by the adrenal zona glomerulosa. Although it has been suggested by many authors that AII acts by increasing the turnover of inositol-lipids, these studies were mainly focussed on the identity and on the kinetics of appearance of inositol phosphates. The purpose of the present study was to establish a relationship between phospholipase C activation and steroidogenesis in the adrenal cortex. A primary culture of bovine adrenal glomerulosa cells was used. Dose-response curves for receptor occupation, inositol phosphate production and aldosterone secretion were made under the same experimental conditions, on the third day of culture. 125I-[Sar1, Val5, D-Phe8]AII binding to glomerulosa cells was progressively inhibited by increasing concentrations of AII up to 30 nM. Scatchard analyses showed a Kd of 1.9 +/- 1.1 nM and a maximal binding capacity of 49,000 +/- 4,500 receptors/cell (six experiments). Dose-response curves for AII-induced inositol 1,4,5-trisphosphate production showed an EC50 of 0.5 +/- 0.1 nM (five experiments). The threshold dose for AII-induced inositol phosphates was around 0.1 nM and the maximal effect was obtained with 30 nM AII. The AII-stimulated steroidogenesis occurred at a threshold dose around 0.03 nM and the maximal effect was obtained with 10 nM AII with an EC50 of 0.5 +/- 0.1 nM (five experiments). These results support previous suggestions that phospholipase C is involved in the steroidogenic action of angiotensin II.

2,3-Diphosphoglycerate is a nonselective inhibitor of inositol 1,4,5-trisphosphate action and metabolism.

Inositol 1,4,5-trisphosphate (InsP3) is an important second messenger generated from the hydrolysis of phosphatidylinositol 4,5-bisphosphate by phospholipase C in response to Ca2(+)-mobilizing stimuli. InsP3 interacts with specific intracellular receptors and triggers the release of sequestered Ca2+ from an intracellular store. We have looked at the influence of 2,3-diphosphoglycerate on the action and metabolism of InsP3 in the bovine adrenal cortex. 2,3-Diphosphoglycerate blocked InsP3 binding to adrenal cortex microsomes with a half-maximal efficiency of 0.5 mM. Scatchard analyses revealed that 2,3-diphosphoglycerate did not change the maximal capacity of the microsomes, but decreased their binding affinity for InsP3. The Ca2(+)-releasing activity of InsP3 on the same microsomal preparation was monitored with the fluorescent indicator, Fura-2. 2,3-Diphosphoglycerate blocked this activity with a half-maximal efficiency of 2 mM. The effect of 2,3-diphosphoglycerate could be overcome by supramaximal doses of InsP3, indicating a competitive inhibitory effect. The activity of InsP3 phosphatase from bovine adrenal cortex microsomes was also studied. 2,3-Diphosphoglycerate inhibited the activity of the phosphatase with a half-maximal efficiency of 0.3 mM. Lineweaver-Burke plots revealed that this effect was competitive. Finally, 2,3-diphosphoglycerate was also able to inhibit the activity of a partially purified preparation of InsP3 kinase from bovine adrenal cortex cytosol. The half-maximal dose was around 10 mM and the Lineweaver-Burke plot showed that the inhibition was competitive. These results show that 2,3-diphosphoglycerate can be considered as a structural analog of InsP3. Its inhibitory effects, however, are not selective enough to use it as an InsP3 protective agent in Ca2(+)-mobilization studies.

Interaction of benzene 1,2,4-trisphosphate with inositol 1,4,5-trisphosphate receptor and metabolizing enzymes.

In a wide variety of cells, inositol 1,4,5-trisphosphate (InsP3) is an important second messenger involved in the regulation of intracellular Ca2+ concentration. InsP3 interacts with specific receptors and triggers the release of sequestered Ca2+ from an internal store. We have synthesized a structural analogue of InsP3 by phosphorylation of the free hydroxyl groups of 1,2,4-benzenetriol with dibenzylphosphorochloridate. The product benzene 1,2,4-trisphosphate (BzP3) was shown to interact with InsP3 receptor and InsP3 metabolizing enzymes of bovine adrenal cortex. BzP3 competitively blocked InsP3 binding to adrenal cortex microsomes with a half-maximal efficiency at 34 microM. This affinity was about 10,000 times lower than that of InsP3 for its receptor. The Ca2+ releasing activity of BzP3 on the same microsomal preparation was monitored with the fluorescent indicator fura-2. BzP3 had no agonistic effect on this activity but it was able to inhibit InsP3-induced Ca2+ release in a dose-dependent manner. The activity of InsP3 phosphatase was also studied. BzP3 inhibited the activity of the phosphatase with a half-maximal efficiency of 32 microM. BzP3 was also able to inhibit the activity of the cytosolic InsP3 kinase with a half-maximal efficiency of 6.1 microM. These results show that BzP3 is interacting with the three specific recognition sites for InsP3 in the bovine adrenal cortex. The inhibitory effect of this compound is relatively more potent on the metabolizing enzymes than on the Ca(2+)-mobilizing receptor.

Interaction of polyanions with the recognition sites for inositol 1,4,5-trisphosphate in the bovine adrenal cortex.

Inositol 1,4,5-trisphosphate (InsP3) serves as a second messenger for Ca2+ mobilization in a wide variety of cells. InsP3 activates a specific receptor/channel located on an internal Ca2+ store. Because heparin has already been shown to block the action of InsP3, we have looked at the influence of other polyanions (dextran sulfate and polyvinyl sulfate) on the action and metabolism of InsP3 in the bovine adrenal cortex. Polyvinyl sulfate blocked InsP3 binding to adrenal cortex microsomes with a half-maximal efficiency of 250 nM. Scatchard analyses revealed that this effect was not competitive. The Ca2+ releasing activity of InsP3 on the same microsomal preparation was monitored with the fluorescent indicator, fura-2. Polyvinyl sulfate blocked this activity with a half-maximal efficiency of 80 nM. The effect of polyvinyl sulfate could not be overcome by supramaximal doses of InsP3, suggesting a non-competitive inhibitory effect. The activity of InsP3 phosphatase from bovine adrenal cortex microsomes was also studied. Polyvinyl sulfate inhibited the activity of the phosphatase with a half-maximal efficiency of 5 microM. Lineweaver-Burk plots revealed that this effect was not competitive. Polyvinyl sulfate was able to inhibit the activity of InsP3 kinase from bovine adrenal cortex cytosol. The half-maximal dose was 15 nM and the Lineweaver-Burk analysis showed that the inhibition was not competitive. The effect of dextran sulfate 5000 (DS-5000) on these activities was also studied. DS-5000 inhibited in a competitive manner the binding of InsP3 to its receptor (IC50 of 34 microM), the release of Ca2+ induced by InsP3 (IC50 of 6.5 microM) and the activity of InsP3 phosphatase (IC50 of 57 microM).(ABSTRACT TRUNCATED AT 250 WORDS)