Biomedical subjects

J R Dave

Publications and source records attributed to J R Dave.

At least 37 records · Page 2Linked to original sources

PubMed1997

Induction of proto-oncogenes during 3-deazaadenosine-stimulated differentiation of 3T3-L1 fibroblasts to adipocytes: mimicry of insulin action.

3-Deazaadenosine (DZA) mimicked the molecular action of insulin in the induction of 3T3-L1 fibroblasts to differentiate into adipocytes. The molecular effects of DZA were compared with insulin, which served as a positive control, on the expression of proto-oncogenes during the initial stage of differentiation of 3T3-L1 fibroblasts. Treatment of confluent 3T3-L1 fibroblasts with DZA or insulin produced a rapid but-transient expression of mRNA for proto-oncogenes c-fos and c-jun within 30-60 min. The mRNA of c-myc increased for 2 h and then decreased 4 h after treatment. Electrophoretic mobility shift assays showed a heightened increase in the appearance of transcription factors AP-1 and AP-2. The increase was detectable as early as 1 h after the treatment with either DZA or insulin and was maintained for 6 h. 3T3-L1 cells stably transfected with the promotor of c-fos linked to a CAT reporter gene showed an increase in CAT activity in response to DZA in a time- and dose-dependent manner. In cells stably transfected with antisense c-fos, neither DZA nor insulin was able to induce a differentiation response. The early transcription of c-fos, c-jun, and c-myc proto-oncogenes and the increased expression of transcription factors AP-1 and AP-2, induced by DZA and insulin, appear to be crucial events in the differentiation of the 3T3-L1 fibroblasts to adipocytes.

3T3 Cells↗

PubMed1995

Phospholipase A2-induced neurotoxicity in vitro and in vivo in rats.

The present study evaluated the neurotoxic potential of phospholipase A2 (PLA2) in in vitro (primary neuronal cultures) and in vivo (EEG and behavior) rat models of CNS excitability. In vitro, PLA2 (0.0038-5.8 nM) or melittin (a potent activator of endogenous PLA2; 100-5000 nM), were highly neurotoxic, causing approximately 500 units/ml LDH release. The neurotoxic EC50s for PLA2 and melittin were 1.8 (1.4-2.3) and 848 (501-1280) nM, respectively. Neurotoxic concentrations of PLA2 stimulated neuronal release of [3H]AA. Preliminary in vitro experiments evaluating changes in neuronal calcium flux indicated that PLA2 caused transient, and melittin sustained, increases in [Ca2+]i. In vivo, PLA2 (0.5-5 micrograms i.c.v.) or melittin (2.5-20 micrograms i.c.v.) produced nonconvulsive EEG seizures, which generalized to status epilepticus. While the onset of seizure development was markedly delayed for PLA2 (1.5-4.5 h), the seizure inducing effects of melittin were evident within 3.5 +/- 0.2 min and more severe. Both PLA2 and melittin were lethal, exhibiting LD50s of 0.62 micrograms and 8.4 micrograms, respectively. Pretreatment with (+)-MK801 (5 micrograms, i.c.v.) significantly attenuated melittin, but not PLA2, in vivo neurotoxicity. PLA2 induced neuropathology in surviving rats revealed extensive cortical and subcortical injury to forebrain neurons and fibre pathways. Collectively, these results demonstrate the potent neurotoxic potential of PLA2, the delayed clinical nature of its in vivo neurotoxicity and the applicability of these model systems to future studies on mechanisms of PLA2 neurotoxicity and the development of potential PLA2 antagonists.

Animals↗

PubMed1994

Regional changes in c-fos mRNA in rat brain after i.v. or i.c.v. NMDA injections.

These studies were undertaken to determine if convulsant doses of i.c.v. vs i.v. administered NMDA exhibit differential specificity for anatomical regions of the brain in stimulating c-fos. In rats i.c.v. or i.v. NMDA produced behaviorally similar clonic (popcorn) convulsions associated with transient increases in c-fos mRNA in different brain areas. Transcription of c-fos mRNA peaked at 30 min post-treatment regardless of the route of administration. However, the route of administration clearly influenced the anatomical specificity of the NMDA-induced c-fos mRNA changes. For example, following i.c.v. administration maximal stimulation in c-fos mRNA was measured in the cerebellum. In contrast, i.v. NMDA produced maximal c-fos mRNA stimulation in the cerebral cortex. Our results demonstrate that NMDA has differential anatomical specificity for molecular signaling in rat brain and suggest that the route of NMDA administration may influence its pathophysiological response.

Animals↗

PubMed1991

Chronic ethanol ingestion decreases vasopressin mRNA in hypothalamic and extrahypothalamic nuclei of mouse brain.

Endogenous arginine vasopressin was previously shown to modulate the rate of loss of functional (CNS) tolerance to ethanol, suggesting that chronic ethanol ingestion might alter vasopressin synthesis and/or release. Since extrahypothalamic vasopressin is believed to be involved in the CNS effects of the peptide, we determined the effect of ethanol on vasopressin mRNA in the bed nucleus of the stria terminalis (BST), as well as in several hypothalamic nuclei. Chronic ethanol ingestion, that produced functional tolerance and physical dependence in mice, resulted in decreased vasopressin mRNA levels in all areas examined. In contrast, as expected, dehydration resulted in increases in vasopressin mRNA in the BST and in all hypothalamic nuclei except the suprachiasmatic nucleus. In the BST, both ethanol ingestion and dehydration affected cells in the central region of the nucleus, while cells in the caudal portion were only affected by ethanol treatment. The results indicate that chronic ethanol ingestion generally reduces the synthesis of vasopressin, and that increased vasopressin synthesis is not necessary in order for the peptide to affect ethanol tolerance.

Animals↗

PubMed1991

Chronic ethanol exposure uncouples vasopressin synthesis and secretion in rats.

To assess the chronic effect of ethanol on vasopressin release and synthesis, hypothalamic vasopressin mRNA, plasma vasopressin levels and plasma osmolality were measured in control rats and rats exposed chronically to ethanol by vapor inhalation for 8 days. The level of hypothalamic vasopressin mRNA (vasopressin synthesis) was unchanged or significantly decreased in ethanol-treated rats, even when these animals displayed increased plasma osmolality and vasopressin levels. The results suggest that chronic ethanol exposure produces a down-regulation of vasopressin synthesis and/or a disruption of vasopressin synthesis-secretion coupling. These findings may have important implications for evaluation of the hydration state of chronic alcoholics.

Animals↗

PubMed1990

Hypothalamic vasopressin mRNA levels in mice are decreased after chronic ethanol ingestion.

Vasopressin mRNA levels in the supraoptic and paraventricular nuclei of the hypothalamus, measured by in situ hybridization with a 35S-labeled RNA probe, were decreased by nearly 50% in C57BL/6NCR mice that had ingested an ethanol-containing diet for 7 days, and were tolerant to and physically dependent on ethanol. At 24 h after withdrawal, vasopressin mRNA levels in the supraoptic nucleus were still decreased, while levels in the paraventricular nucleus returned toward control values. Although plasma osmolality was increased in ethanol-fed mice, there was no increase in plasma vasopressin levels, possibly as a result of the effect of chronic ethanol ingestion to decrease vasopressin synthesis. In contrast, in mice that were dehydrated, but not fed ethanol, plasma osmolality, plasma vasopressin levels, and hypothalamic vasopressin mRNA all increased, as expected. The data suggest that chronic ethanol ingestion interferes with the synthesis and secretion of vasopressin, and may result in the reduced ability of an individual to respond to physiological stimuli for vasopressin secretion.

Animals↗

PubMed1990

Reduction of complement activation during bypass by prime manipulation.

Complement activation is believed to be of importance in the development of complications arising after cardiopulmonary bypass. The effect on complement activation of priming the extracorporeal circuit with crystalloid alone, crystalloid plus albumin, or crystalloid plus the plasma expander polygeline was assessed in 36 patients undergoing coronary artery operations with cardiopulmonary bypass using a bubble oxygenator. Activation of the alternative and common complement pathways was monitored before, during, and after the bypass period by measuring concentrations of factor B and its fragment Ba and C3 and its fragment C3d. Complement activation occurred in all three groups of patients, with no difference between the crystalloid and crystalloid-albumin groups. In contrast, Ba fragment concentrations were persistently and significantly lower during and after bypass in the polygeline group, denoting reduced complement activation. C3d levels also showed a tendency to be lower in this group. Our results indicate that addition of polygeline to the priming solution reduces complement activation. Because complement activation is associated with morbidity after cardiopulmonary bypass, addition of polygeline to the priming solution may offer an inexpensive method of reducing morbidity after cardiopulmonary bypass.

Albumins↗

PubMed1990

Arginine vasopressin induces the expression of c-fos in the mouse septum and hippocampus.

Arginine vasopressin is a neuropeptide that has been shown to modulate functional ethanol tolerance and memory processes. These actions of vasopressin in the CNS have been shown by us and others to be mediated by V1 receptors. Intracerebroventricular injection of vasopressin in mice resulted in a substantial increase in mRNA for the proto-oncogene c-fos in septum and hippocampus, but no increase in cerebral cortex. A V1-selective agonist also increased septal c-fos mRNA levels, while a V2-selective agonist was less effective. Similarly, the response to vasopressin was more effectively blocked by a V1- than a V2-selective antagonist. These results indicate that vasopressin acts specifically at V1 receptors in mouse septum and hippocampus to increase c-fos mRNA. The vasopressin metabolite, AVP(4-9), also increased c-fos mRNA levels in septum and hippocampus, while the response to oxytocin, which has different effects from vasopressin on memory and tolerance, was greater in hippocampus than in septum. Nerve growth factor, in contrast to the other peptides, had a more pronounced effect on c-fos mRNA levels in cerebral cortex than in the other brain areas. Increased c-fos expression has been hypothesized to play a role in neuroadaptation, and these results suggest that modulation of septal c-fos expression could be important for vasopressin effects on ethanol tolerance and/or memory.

Animals↗

PubMed1990

Complement activation before, during and after cardiopulmonary bypass.

Plasma levels of the complement parent molecules C3, C4, and factor B and their split products, C3d, C4d, and Ba were measured in 12 patients undergoing cardiopulmonary bypass for coronary artery surgery. Alternative and common complement pathway activation, demonstrated by statistically significant rising levels of Ba (P less than 0.05), and C3d (P less than 0.05) and by elevated Ba:B (P less than 0.05) and C3d:C3 (P less than 0.05) ratios were found before the institution of cardiopulmonary bypass but following heparin administration suggesting that heparin may itself initiate alternative pathway activation. In addition, significant depletion of parent complement components and elevation of split product concentrations was seen during bypass suggesting classical and alternate pathway activation (P less than 0.01). This study clarifies the pathways of complement activation during bypass and presents evidence that heparin administration may initially activate the complement cascade.

Cardiopulmonary Bypass↗

PubMed1990

The role of arginine vasopressin in alcohol tolerance.

Administration of the neuropeptide, arginine vasopressin, to animals that have acquired functional tolerance to ethanol will maintain such tolerance, even in the absence of further ethanol ingestion by the animals. In mice, this action of the peptide is mediated by central nervous system V1 receptors and requires intact brain noradrenergic systems. Autoradiographic studies have shown that some V1 receptors are localized presynaptically on catecholaminergic neuronal terminals in the mouse lateral septum, suggesting that vasopressin may act via modulation of catecholamine release. In addition, vasopressin has been found to increase mRNA levels for the proto-oncogene, c-fos, in septum and hippocampus, possibly by an action at postsynaptic receptors. Expression of c-fos, which has been hypothesized to play a role in central nervous system neuroadaptation, could transform short-term actions of vasopressin into long-term effects on ethanol tolerance. Studies with vasopressin antagonists indicate that the endogenous peptide influences tolerance, and therefore the effect of chronic ethanol ingestion on vasopressin synthesis and release was studied. In mice and rats, hypothalamic vasopressin mRNA is decreased by chronic ethanol exposure, although effects on plasma vasopressin levels differ in the two species. The effect of ethanol on extrahypothalamic vasopressin synthesis in brain is under investigation. The results suggest mechanisms by which vasopressin can produce long-term changes in central nervous system function, and provide evidence for a disturbance of vasopressin regulation during chronic ethanol ingestion.

Alcohol Drinking↗

PubMed1990

Ethanol withdrawal seizures produce increased c-fos mRNA in mouse brain.

mRNA levels for the protooncogene c-fos, measured by Northern blot analysis, were greatly increased in brains of mice undergoing ethanol withdrawal seizures. This increase was transient (levels were increased at the time of the seizure and returned to normal by 24 hr or less after seizure) and was larger in hippocampus (40-fold) than in cerebral cortex (10-fold) or in cerebellum (6-fold). In mice that were fed ethanol chronically and withdrawn but that did not undergo overt withdrawal seizures, c-fos mRNA levels were not significantly increased. The findings with ethanol withdrawal seizures are similar in many respects to results of earlier studies with chemically induced seizures or kindling, which had led to the suggestion that c-fos expression may play a role in neuronal adaptation. The development of ethanol withdrawal seizures has been likened to kindling, and there is evidence indicating that ethanol withdrawal symptoms become more severe after repeated episodes of withdrawal. The present data support the hypothesis that this phenomenon may involve ethanol withdrawal seizure-induced increases in c-fos expression in various brain areas.

Animals↗

PubMed1989

Rubella virus: mechanism of attenuation in the vaccine strain (HPV77).

The vaccine type (HPV77 strain) of rubella virus replicates slower and manifests a delayed appearance of cytopathic effect in Vero-76 cells as compared to wild-type virus (M33). The change in cytopathic effect coincides with the delayed appearance of both genomic and subgenomic RNA as well as viral structural proteins in the cell. The delay in the appearance of the viral proteins in the cells was also evident when the cells infected with the vaccine-type virus were treated with the lysosomotropic agent such as chloroquine. Binding studies using [35S]methionine-labeled virus showed that the vaccine-type virus bound to the cells poorly and the binding was not completely competed out with the cold virus.

Animals↗

PubMed1988

Regulation of vasopressin and oxytocin synthesis in anterior pituitary and peripheral tissues.

Recent studies have demonstrated the presence of immunoreactive oxytocin (OT) and vasopressin (VP), OT and VP receptors and physiological functions for these two hormones in a variety of peripheral tissues, including anterior pituitary gland. The objectives of this study were to determine if (i) OT and VP genes are expressed in rat testis and anterior pituitary gland and (ii) if osmotic stimulation known to modify the regulation of OT and VP genes in hypothalamus, would modify the expression of these genes in rat testis and anterior pituitary gland. Using oligonucleotide probes (courtesy of Drs. M. Brownstein and W. Scott Young, NIMH) corresponding to the VP gene or OT gene and specific fractions of human OT and VP genes (courtesy of Dr. J. Battey, NCI) subcloned in the pGEM-3 riboprobe system, and Northern blot and slot blot techniques, OT and VP mRNAs were found in rat testis and anterior pituitary gland. When adult male rats (SD) were either deprived of drinking water or offered 2% salt solution as a sole source of drinking fluid for 72 hrs, both OT and VP mRNA levels were increased in hypothalamus, anterior pituitary gland and testis. Our data suggest that testis and anterior pituitary gland could also be sites of synthesis of OT and VP and that the same stimulus may regulate these genes in various tissues.

Animals↗

PubMed1987

Barium distinguishes separate calcium targets for synthesis and secretion of peptides in neuroendocrine cells.

The effect of barium and potassium on the secretion and biosynthesis of enkephalin in bovine chromaffin cells, and prolactin and beta-endorphin in rat anterior pituitary cells, was examined to determine whether calcium-dependent secretion and biosynthesis are mediated by the same or by different calcium targets within the neuroendocrine cell. In the presence of 1.8 mM calcium, barium and potassium stimulated the secretion of all three peptides over 30 min, and increased the levels of proenkephalin and prolactin mRNA in 24 hr. These effects were inhibited by the calcium channel blocker D600. When the extracellular calcium concentration was lowered to 0.1 mM or less, secretion elicited by potassium was blocked, whereas secretion elicited by barium was enhanced, indicating that barium wholly substitutes for extracellular calcium in mediating peptide secretion. On the other hand, stimulation of proenkephalin and prolactin mRNA by both potassium and barium was inhibited when the extracellular calcium concentration was reduced. We conclude that calcium acts at two different intracellular targets to activate secretion versus biosynthesis of both enkephalin and prolactin. This appears to be the first report in which two different calcium-dependent processes in the intact cell are distinguished by a calcium ion agonist. Calcium-dependent processes such as protein phosphorylation, protein translocation, and enzyme activation may thus be related to events in the intact cell such as peptide synthesis and secretion on the basis of selective stimulation by barium.

Animals↗

PubMed1987

Anterior pituitary gland modulates the release of atrial natriuretic peptides from cardiac atria.

Myocytes in the atria contain a prohormone that gives rise to atrial natriuretic peptides (ANP), which have intrinsic hemodynamic regulatory activity. Little is known about mechanisms regulating ANP release. In rats with indwelling catheters, acute blood volume expansion with 5% (wt/vol) dextrose increases the amount of circulating immunoreactive ANP by a factor of 2-3, as determined by radioimmunoassay. Pithing, which both removes neurogenic influences and interrupts humoral influences of the brain and pituitary gland on the heart, completely blocked stimulus-induced release of ANP. Because our studies using pharmacological blockade of the autonomic nervous system had suggested that neurogenic mechanisms do not play a major role in ANP release, we sought a humoral mechanism involved in ANP secretion. Basal and stimulated release of ANP were significantly blunted in hypophysectomized rats (8 days after operation) but were completely restored when the resected anterior pituitary was reimplanted under the kidney capsule. This suggests that hormones of anterior pituitary origin are required for ANP secretion in response to acute volume loading.

Adrenalectomy↗

PubMed1987

Calcium-independent and calcium-dependent mechanisms regulate corticotropin-releasing factor-stimulated proopiomelanocortin peptide secretion and messenger ribonucleic acid production.

We have evaluated the role of calcium in basal and secretagogue-stimulated release of beta-endorphin and PRL and the levels of their respective mRNAs in primary cultured rat anterior pituitary cells. Treatment of anterior pituitary cells with the calcium channel blocker methoxyverapamil (D600; 10 microM) or with calcium-free medium for 1 h did not alter the basal release of beta-endorphin and only partially blocked CRF (10 nM)-stimulated beta-endorphin release. In contrast to these effects of D600 or calcium-free medium on corticotrophs, both of these test conditions decreased basal secretion of PRL from lactotrophs by 50-70% and completely blocked forskolin (10 microM)-stimulated PRL secretion. Although omission of calcium from the culture medium caused a 50% decrease in basal levels of both proopiomelanocortin (POMC) and PRL mRNA, treatment of cells with D600 did not significantly alter the basal levels of POMC or PRL mRNA. Treatment of cells with D600 partially blocked CRF-stimulated elevation of POMC mRNA and forskolin-stimulated elevation of PRL mRNA. The calcium agonist barium (1 mM) produced a 2-fold increase in both beta-endorphin and PRL release, which was blocked by D600. Treatment of cells with barium had no effect on POMC mRNA levels, but increased PRL mRNA levels. D600 treatment of cells partially blocked barium-stimulated PRL mRNA levels. These findings demonstrate a calcium-dependent as well as a calcium-independent component of CRF-stimulated beta-endorphin secretion and CRF-stimulated POMC mRNA elevation. In contrast, PRL secretion and biosynthesis appear to be totally calcium-dependent processes.

Animals↗

PubMed1986

Circulating atrial natriuretic peptides in conscious rats: regulation of release by multiple factors.

Cardiocytes in the atria contain a prohormone that gives rise to atrial natriuretic peptides (ANP's), which have intrinsic hemodynamic regulatory activity. The distribution of ANP's in the brain suggests the involvement of these peptides in central cardiovascular regulation. In conscious rats with chronic indwelling catheters, volume loading with isotonic saline or glucose increased the amount of circulating immunoreactive ANP's by a factor of 4 to 5, as determined by radioimmunoassay. Hyperosmotic challenge with a hypertonic NaCl solution or anesthesia with halothane caused similar increases in plasma ANP's. Results obtained with the denervated-heart preparation indicate that neuronal influences are important in the release of ANP's induced by volume loading. As judged from reversed-phase high-performance liquid chromatography of extracted plasma and radioimmunoassay of collected fractions, the circulating physiologically important ANP's in the conscious rodent appear to be alpha-rANP(5-28) (atriopeptin III) and either alpha-rANP(3-28) [ANF(8-33)] or alpha-rANP(1-28) (ANF).

Anesthesia↗

PubMed1986

Demonstration of corticotropin-releasing factor binding sites on human and rat erythrocyte membranes and their modulation by chronic ethanol treatment in rats.

In a previous study we reported the presence of specific corticotropin-releasing factor (CRF) binding sites in peripheral tissues of the rat (Endocrinology, 116, 2152, 1985). Using 125I-labeled rat or human CRF, specific CRF binding sites were identified on rat and human erythrocytes, but not on lymphocytes or platelets. Furthermore, identical CRF binding was observed in the presence of intact erythrocytes or lysed erythrocyte membranes. Maximal binding of 125I-CRF occurred within 25 min at 4 degrees C and was saturable. Scatchard analysis of CRF binding to erythrocyte membranes revealed the existence of a single class of binding site. Chronic exposure of rats to ethanol vapor, known to lower specific CRF binding to pituitary tissue by 35%, also decreased 125I-rat CRF binding to erythrocyte membranes by approximately 45%, which was due to a decrease in the number of CRF binding sites. The parallel decrease of CRF binding to rat-erythrocyte and pituitary membranes following chronic ethanol treatment suggests that CRF binding to erythrocyte and pituitary membranes is modulated in a similar direction, which further suggests that the determination of CRF binding to erythrocytes may provide an important clinical tool to indirectly assess CRF-receptor levels in the pituitary gland and thereby enhance our understanding of ethanol-induced disorders of the hypothalamic-pituitary-adrenal axis in patients.

Animals↗