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S Rattan

Publications and source records attributed to S Rattan.

At least 37 records · Page 2Linked to original sources
PubMed1997

L-citrulline recycling in opossum internal anal sphincter relaxation by nonadrenergic, noncholinergic nerve stimulation.

BACKGROUND & AIMS: L-citrulline formed stoichiometrically along with nitric oxide (1:1) from L-arginine may be enzymatically converted to L-arginine. The possibility of L-citrulline recycling in the maintenance of nitrergic neurotransmission in the opossum internal anal sphincter (IAS) smooth muscle strips was investigated. METHODS: Responses to nonadrenergic, noncholinergic (NANC) nerve stimulation by electrical field stimulation (EFS) (either short-train or continuous stimulation) on the basal IAS tension were recorded before and after the NO synthase inhibitor N(omega)-nitro-L-arginine (L-NNA), L-NNA plus L-citrulline, or L-arginine. During continuous EFS, when the basal IAS tone after the initial relaxation had recovered to almost pre-EFS levels, the effects of L-citrulline or L-arginine were examined before and after L-glutamine, which is a putative blocker of L-citrulline uptake. RESULTS: Inhibition of NANC nerve-mediated IAS relaxation by L-NNA was reversed by L-citrulline as well as L-arginine. L-Citrulline and L-arginine caused concentration-dependent relaxation of the IAS tone recovered during the prolonged EFS. L-Glutamine blocked the responses of L-citrulline but not of L-arginine. Furthermore, L-glutamine increased the speed of recovery of IAS tone during continuous EFS. CONCLUSIONS: L-citrulline recycling may be responsible for the maintenance of IAS relaxation during frequent short-train and prolonged NANC nerve stimulation.

Anal Canal↗
PubMed1997

Regulation of inducible and neuronal nitric oxide synthase gene expression by interferon-gamma and VIP.

The studies examined the regulation of inducible and neuronal nitric oxide synthases (iNOS and nNOS, respectively) in the rat brain, stomach, rectum, and spleen. Relative expression of iNOS and nNOS mRNAs was quantified by the sensitive method of polymerase amplification reactions. The NOS proteins were determined by Western blot, using specific antibodies. Highest levels of nNOS and iNOS mRNAs were expressed in the rat brain and spleen, respectively. Furthermore, both nNOS and iNOS were expressed in the stomach and rectum. Interestingly, treatment of tissues with lipopolysaccharides or cytokine interferon-gamma (IFN-gamma) induced the expression of iNOS and decreased that of nNOS, representing a shift from one isoform to the other. When the tissues were treated with IFN-gamma followed by vasoactive intestinal polypeptide (VIP), the induction of iNOS was reduced by VIP. The changes in iNOS and nNOS expression at the transcriptional level corresponded to those at the translational level. The data suggest a regulatory role of IFN-gamma and VIP in the expression iNOS and nNOS and a counterregulation of iNOS and nNOS in rat tissues.

Animals↗
PubMed1997

Neuronal NOS gene expression in gastrointestinal myenteric neurons and smooth muscle cells.

Nitric oxide synthase (NOS) has been characterized in different tissues, and its localization has been suggested in different neuronal tissues, including the myenteric neurons and other nonneuronal cells. The present study examined the distribution of the neuronal NOS (nNOS) mRNA in different tissues of the opossum gastrointestinal tract, internal anal sphincter (IAS) smooth muscle cells, and myenteric neurons using slot-blot and Northern blot hybridization techniques with a specific rat brain nNOS cDNA probe. Significant levels of nNOS gene expression were found in both smooth muscle cells and myenteric neurons of the opossum IAS. This finding was confirmed by reverse transcriptase-polymerase chain reaction analysis of the RNA obtained from cultured opossum IAS smooth muscle cells and myenteric neurons and also from human intestinal smooth muscle and neuroblastoma cell lines. Pyloric sphincter had the highest level of nNOS gene expression compared with other gastrointestinal tissues. There was no significant difference in the nNOS gene expression between other sphincteric and nonsphincteric tissues examined. The present study shows the presence of nNOS gene expression in both neurons and smooth muscle cells. The higher levels of nNOS gene expression in the pyloric sphincter compared with other tissues may have pathophysiological significance in some disease conditions.

Animals↗
PubMed1997

L-arginine deficiency causes suppression of nonadrenergic noncholinergic nerve-mediated smooth muscle relaxation: role of L-citrulline recycling.

Studies were performed on the internal anal sphincter (IAS) smooth muscle strips obtained from opossums (Didelphis virginiana). Isometric tension and L-arginine levels of the tissues were measured under basal conditions, in the presence of electrical field stimulation (EFS) and after treatment with different concentrations of arginase. For the nonadrenergic noncholinergic nerve stimulation, short trains (4 sec) as well as continuous EFS were used. During continuous EFS, after the initial IAS relaxation, the response began to fade within several min to approximately 80% recovery of the basal tone. We also examined the influence of L-arginine and L-citrulline on these responses. For some studies, the tissues were pretreated with L-glutamine (an inhibitor of L-citrulline uptake), L-glutamate or N(G)-hydroxy-L-arginine (an inhibitor of arginase). Interestingly, the basal levels of L-arginine were found to be significantly higher in the IAS (tonic smooth muscle) than in the rectal (phasic smooth muscle) smooth muscle. Arginase caused a concentration-dependent attenuation of the IAS relaxation caused by EFS. L-Citrulline and L-arginine were equipotent in reversing the attenuation. Both arginase (60 min pretreatment) and continuous EFS (tissues collected at the time of maximal recovery of the basal IAS tone after the initial relaxation) caused significant decreases in L-arginine levels. The decreases in the levels of L-arginine were restored by the exogenous administration of either L-arginine or L-citrulline. The restoration of L-arginine levels by L-citrulline but not by L-arginine was selectively blocked by L-glutamine. Furthermore, the IAS relaxation, attenuated by arginase was unaffected by L-glutamine but was restored by N-hydroxy-L-arginine pretreatment. The studies suggest that L-citrulline-L-arginine recycling may play a significant role in the maintenance of IAS relaxation in response to nonadrenergic noncholinergic nerve stimulation.

Anal Canal↗
PubMed1997

Excitatory and inhibitory actions of pituitary adenylate cyclase-activating peptide (PACAP) in the internal anal sphincter smooth muscle: sites of actions.

Unlike its effects on the rest of the GI tract, the effects of pituitary adenylate cyclase-activating peptide (PACAP) on the internal anal sphincter (IAS) are not known. We examined the actions of PACAP-38 (here PACAP) and PACAP-27 on the basal IAS tone of circular smooth muscle strips before and after the administration of different neurohumoral antagonists. PACAP caused a concentration-dependent fall in the basal tone of the IAS. Interestingly, however, at higher concentrations, PACAP caused a biphasic response: an initial contraction followed by a relaxation. Both the contractile and the relaxant responses were insensitive to atropine, guanethidine, apamin or tetrodotoxin. Both the contractile and the relaxant effects were inhibited by PACAP 6-38 (a selective antagonist of PACAP), vasoactive intestinal polypeptide 10-28 (a vasoactive intestinal polypeptide antagonist) and PACAP tachyphylaxis. The nitric oxide synthase inhibitor Nomega-nitro-L-arginine attenuated the inhibitory but not the excitatory effect of PACAP. Conversely, the contractile but not the relaxant effect of PACAP on the IAS was nearly obliterated by the substance P antagonist spantide. The N-type Ca++-channel blocker omega-conotoxin caused significant suppression of both the contractile and the inhibitory actions of PACAP. We conclude that in the IAS, PACAP has a dual effect: a contraction followed by a relaxation. The contraction of IAS by PACAP is speculated to occur via the activation of PACAP receptor at the substance P-containing nerve terminals. PACAP-induced IAS relaxation, on the other hand, appears to be mediated in large part by its direct action at the smooth muscle cells and in part by its action at the nerve terminals of the myenteric inhibitory neurons.

Anal Canal↗
PubMed1996

Evidence for VIP-induced increase in NO production in myenteric neurons of opossum internal anal sphincter.

A significant interaction between vasoactive intestinal polypeptide (VIP) and nitric oxide (NO) has been reported in neurotransmission of the gastrointestinal tract, including the internal anal sphincter (IAS). The exact site of this NO release from the IAS in response to VIP is not known. Studies were carried out to determine the site of this VIP-induced NO release in opossum IAS. NO synthase (NOS) activity was quantitated by determining L--3H-citrulline production from L[3H]arginine in isolated myenteric ganglia and smooth muscle cells of the IAS. L-[3H]citrulline production was determined before and after treatment with either the ganglionic stimulant 1,1-dimethyl-4-phenylpiperazinium iodide (DMPP), VIP, or peptide histidine isoleucine (PHI) in the absence and presence of the neurotoxin tetrodotoxin and the NOS inhibitor NG-nitro-L-arginine (L-NNA). Smooth muscle cells and ganglia were preloaded with L-[3H]arginine for 5 min and treated with VIP for 1 and 5 min. DMPP and VIP caused a significant increase in L-[3H]citrulline formation in myenteric ganglia at both time periods, whereas in smooth muscle cells there was a moderate but significant increase in L-[3H]citrulline production only at 5 min of VIP treatment. VIP-induced relaxation of isolated smooth muscle cells of the IAS was not affected by L-NNA. The increase in NOS activity of myenteric ganglia by DMPP and VIP was sensitive to neurotoxin and the NOS inhibitor. The data suggest that the increase in NO production in response to VIP in the IAS occurs mainly from the myenteric neurons, with some contribution from the smooth muscle cells.

Anal Canal↗
PubMed1996

Heme oxygenase inhibitor zinc protoporphyrin IX causes an activation of nitric oxide synthase in the rabbit internal anal sphincter.

The studies were performed in the rabbit internal anal sphincter (IAS) smooth muscle strips to examine the influence of the heme oxygenase inhibitor, [zinc protporphyrin (ZnPP IX)], on basal tone. ZnPP IX produced a concentration-dependent fall in the basal tone and was the focus of our investigation. To examine the mechanism of the fall in IAS tone by ZnPP IX, the effect of different concentrations of ZnPP IX on basal IAS tone and the release of nitric oxide were examined before and after the neurotoxin tetrodotoxin and various nitric oxide synthase (NOS) inhibitors. The inhibitory effect of ZnPP IX was blocked by the NOS inhibitors L-NG-nitroarginine, NG-monomethyl-L- arginine and L-N5-(1-iminoethyl)ornithine and the neurotoxin TTX. The fall in IAS tension by ZnPP IX was accompanied by a release of NO. ZnPP IX(1 x 10(-3)M) caused a fall in IAS tension of 43.7% which was reduced to 16.5% in the presence of L-NG-nitroarginine (1 x 10(-4)M). Furthermore, the fall in IAS tone in the presence of ZnPP IX was restored both by the NOS inhibitors and tetrodotoxin. The basal release of nitric oxide in these experiments was 0.50 +/- 0.07 nmol and in the presence of ZnPP IX (1 x 10(-3)M), it increased to 1.72 +/- 0.28 nmol (more than a 3-fold increase). Thus the fall in the basal IAS tone by ZnPP IX was associated with a release of NO from the myenteric neurons as these effects were significantly blocked by the NOS inhibitors and tetrodotoxin. We conclude that in the rabbit IAS, ZnPP IX causes a fall in the basal IAS tension by the activation of NOS in myenteric neurons. We speculate that in the resting state, the heme oxygenase pathway exerts important counter-regulatory effects on the NOS pathway and when blocked (e.g., by ZnPP IX), the underlying NOS pathway is unmasked leading to a massive and prolonged release of NO. The exact significance of this mechanism remains to be determined.

Anal Canal↗
PubMed1995

Definition and refinement of chromosome 11 regions of loss of heterozygosity in breast cancer: identification of a new region at 11q23.3.

Chromosome 11 is frequently altered in several types of human neoplasms. In breast cancer, loss of heterozygosity has been described in two regions of this chromosome, 11p15 and 11q22-23. In this report we have dissected the two regions using high-density polymorphic markers, and have found that there are at least two independent areas of loss of heterozygosity in each region, suggesting that multiple genes on chromosome 11 may be targets of genetic alteration during tumor establishment or progression. The regions defined are: at 11p15, between loci D11S576 and D11S1318 and between D11S988 and D11S1318; at 11q23, between D11S2000 and D11S897 and between D11S528 and D11S990. The narrowing of these regions of loss should facilitate the cloning of the regions in yeast artificial chromosomes to identify the critical tumor suppressor genes.

Base Sequence↗
PubMed1995

Seven megabase yeast artificial chromosome contig at region 11p15: identification of a yeast artificial chromosome spanning the breakpoint of a chromosomal translocation found in a case of Beckwith-Wiedemann syndrome.

Genetic alterations of chromosome region 11p15 have been detected in neoplastic diseases as well as in cancer-predisposing syndromes. The cloning of the entire chromosomal region will be important for the identification and characterization of critical tumor suppressor genes. We have developed a yeast artificial chromosome contig that covers up to 7 Mb of this chromosome band. The most centromeric marker included in the contig is D11S932 and the most telomeric is D11S470. We have developed 18 new STS markers, which have been located in the contig in relation to 16 known markers. One of the yeast artificial chromosome clones was found to span the chromosome 11 breakpoint of the translocation t(11;18), associated with a case of Beckwith-Wiedemann syndrome. Cloning the regions in proximity to this translocation might reveal the presence of a gene altered in association with the development of Beckwith-Wiedemann syndrome.

Base Sequence↗
PubMed1995

Colocalization of NADPH-diaphorase staining and VIP immunoreactivity in neurons in opossum internal anal sphincter.

Nitric oxide and vasoactive intestinal polypeptide (VIP) are important inhibitory neurotransmitters mediating relaxation of the internal anal sphincter. The location and coexistence of these two neurotransmitters in the internal anal sphincter has not been examined. We performed a double-labeling study to examine the coexistence of nitric oxide synthase and VIP in the opossum internal anal sphincter using the NADPH-diaphorase technique which is a histochemical stain for nitric oxide synthase. In perfusion-fixed, frozen-sectioned tissue, VIP-immunoreactive neurons were labeled using immunofluorescence histochemistry. After photographing the VIP-immunoreactive neurons, nitric oxide synthase was labeled using the NADPH-diaphorase technique. Ganglia containing neuronal cell bodies were present in the myenteric plexus for the entire extent of the internal anal sphincter. VIP-immunoreactive and NADPH-diaphorase-positive neurons were present in ganglia in the myenteric as well as the submucosal plexuses. Most of the VIP-immunoreactive neurons were also NADPH-diaphorase positive. VIP and nitric oxide synthase are present and frequently coexist in neurons in the internal anal sphincter of the opossum. These neurons may be an important source of inhibitory innervation mediating the rectoanal reflex-induced relaxation of the sphincter. The demonstration of the coexistence of these two neurotransmitters will be of fundamental importance in unraveling their relationship and interaction in the internal anal sphincter as well as other systems.

Amino Acid Oxidoreductases↗
PubMed1995

In vivo and in vitro influence of human recombinant hemoglobin on esophageal function.

The purpose of the present investigation was to examine the influence of a nitric oxide scavenger, hemoglobin (Hb), on esophageal function. Intraluminal pressures of anesthetized opossums were recorded from lower esophageal sphincter (LES) and 1, 5, and 9 cm above the LES. The influence of a representative Hb-based oxygen carrier was examined on swallowing-induced esophageal peristalsis and LES relaxation. In in vitro studies, LES relaxation and esophageal peristaltic contractions were induced by the activation of nonadrenergic noncholinergic (NANC) neurons by electrical field stimulation (EFS). Hb caused significant impairment in swallowing- and EFS-induced LES relaxation and a significant increase in the speed of esophageal peristalsis. In some experiments, swallowing caused simultaneous contractions in the esophagus following Hb administration. Although Hb completely blocked LES relaxation by NO and significantly attenuated that by NANC nerve stimulation, it had no significant effect on isoproterenol-induced LES relaxations. The data support the role of NO in LES relaxation and esophageal peristalsis. This esophageal model may be important in understanding the influence of NO inhibitors and scavengers in gastrointestinal motility.

Animals↗
PubMed1995

Distribution of VIP binding sites in opossum internal anal sphincter circular smooth muscle.

The main goal of the present study was to determine whether vasoactive intestinal polypeptide (VIP) binds exclusively to smooth muscle plasma membranes or also to neurons. We examined the distribution of VIP binding sites in different fractions and subfractions of smooth muscle membranes prepared from the circular muscle layer of opossum internal anal sphincter (IAS). The markers used for synaptosomal and smooth muscle plasma membranes were [3H]saxitoxin binding and 5' nucleotidase activity, respectively. Fractionation of the membranes was carried out by differential centrifugation of the circular smooth muscle membranes. Further subfractionations were done by sucrose density gradient centrifugation. VIP binding was high in the membrane fractions that were enriched with 5' nucleotidase activity. However, some VIP binding was also found in fractions with high saxitoxin binding. Membranes prepared from isolated smooth muscle cells and myenteric neurons both had high VIP binding. Membranes prepared from isolated smooth muscle cells displayed higher 5' nucleotidase activity, whereas membranes prepared from isolated myenteric plexi had higher saxitoxin binding. In conclusion, in the opossum IAS circular smooth muscle, binding of VIP occurs on both the smooth muscle plasma membranes and the synaptosomal membranes. The presence of VIP binding sites on the smooth muscle plasma membrane supports the role of VIP as an inhibitory neurotransmitter. The significance of binding of the neuropeptide to the neuronal membranes remains to be determined.

5'-Nucleotidase↗
PubMed1995

Human recombinant hemoglobin (rHb1.1) inhibits nonadrenergic noncholinergic (NANC) nerve-mediated relaxation of internal anal sphincter.

Nitric oxide (NO) plays a significant role in the nonadrenergic noncholinergic nerve-mediated relaxation of gastrointestinal smooth muscle. Furthermore, hemoglobin is known to avidly bind NO and inhibit the nonadrenergic noncholinergic nerve-mediated smooth muscle relaxation. The influence of recombinant hemoglobin (rHb1.1) on gastrointestinal smooth muscle relaxation is not known. In this study, we examined the influence of rHb1.1 on the opossum internal anal sphincter (IAS) relaxation in response to electrical field stimulation, nitric oxide, vasoactive intestinal polypeptide, peptide histidine isoleucine and isoproterenol. The IAS smooth muscle strips developed spontaneous tone and exhibited frequency-dependent relaxation in response to electrical field stimulation. rHb1.1 caused concentration-dependent attenuation (30%-85%) of electrical field stimulation-induced IAS relaxation and complete obliteration of IAS relaxation induced by different concentrations of NO. rHb1.1 also caused suppression of the inhibitory effects of vasoactive intestinal polypeptide on IAS. Fall in the IAS tension by peptide histidine isoleucine and by isoproterenol were not modified by rHb1.1. The rHb1.1-induced blockade of neurally mediated IAS relaxations provides further support for the role of NO as an inhibitory neurotransmitter/mediator in the IAS.

Anal Canal↗
PubMed1994

Suppression of tumorigenicity of breast cancer cells by microcell-mediated chromosome transfer: studies on chromosomes 6 and 11.

Development of breast cancer has been associated with deletions at multiple chromosomal regions, including 6q, 11p, and 11q. In this study we analyzed the effects of the introduction of chromosomes 6 and 11 on the cell phenotype of the breast cancer cell lines MDA-MB-231 and MCF-7. Chromosome 6 induced alterations of in vitro growth properties and suppressed tumorigenicity of MDA-MB-231 cells. Spontaneous reduction of the transferred chromosome allowed mapping of the tumor suppressor gene(s) to region 6q21-q23 and/or 6q26-q27. Clones MCF-7/H6 underwent a senescence process that lasted five months. Chromosome 11 had no effect on MDA-MB-231 cells, although it suppressed tumorigenicity of MCF-7 cells. A MCF-7/H11 clone lacking the short arm of the transferred chromosome retained tumorigenicity, however, tumor cell growth was significantly reduced. These results suggest that each chromosomal arm may contain genes important for the suppression of MCF-7 tumorigenic properties.

Breast Neoplasms↗
PubMed1994

Vasoactive intestinal polypeptide gene expression is characteristically higher in opossum gastrointestinal sphincters.

BACKGROUND/AIMS: Vasoactive intestinal polypeptide (VIP) has been suggested to be an inhibitory neurotransmitter in the sphincteric and nonsphincteric smooth muscles of the gut. However, the relative gene expression of VIP in these functionally diverse regions is not known. METHODS: The gastrointestinal smooth muscle sphincters of opossums were excised from the adjoining nonsphincteric smooth muscles. RNAs were isolated and subjected to blot hybridizations with VIP complementary DNA probe. Relative expression of VIP gene was quantitated using the densitometric scanning of the VIP messenger RNA (mRNA) transcripts. The cellular specificity of VIP gene expression was investigated in cultures of neuroblastoma cells and myenteric plexuses and compared with those of the smooth muscle cells. RESULTS: The data showed higher levels of VIP mRNA in the sphincteric than the adjoining nonsphincteric tissues. VIP mRNA were found in significantly higher amounts in the myenteric neurons and neuroblastoma cells than in the smooth muscle cells. CONCLUSIONS: VIP gene expression was significantly higher in the sphincteric smooth muscle regions than in the nonsphincteric regions of the gut. The studies provide further evidence for the role of VIP in neurotransmission of the gut.

Anal Canal↗
PubMed1994

Differential isoactin gene expression in the sphincteric and nonsphincteric gastrointestinal smooth muscles of the opossum.

The sphincteric smooth muscle tissues of the gastrointestinal tract have been shown to possess mechanical properties distinct from the surrounding nonsphincteric smooth muscle tissues. Little is currently known regarding the molecular basis of this differential smooth muscle development and function. Actin is an important contractile protein whose expression has been linked to the normal development and function of smooth muscle tissues. The purpose of this study was to characterize isoactin gene expression in the sphincteric versus nonsphincteric smooth muscle tissues of the gastrointestinal tract. Northern blot analysis was performed on manometrically identified sphincteric and the flanking nonsphincteric smooth muscle tissues of the adult opossum. Quantitative analysis revealed a distinct pattern of isoactin gene expression in the sphincteric versus nonsphincteric smooth muscle tissues of the gut. The sphincteric smooth muscle tissues expressed significantly lower total quantities of isoactin mRNA than their surrounding nonsphincteric smooth muscle tissues. It is hypothesized that these differential patterns of isoactin gene expression may play a significant role in establishing the myogenic potential of the functionally distinct sphincteric and nonsphincteric smooth muscle tissues of the gut.

Actins↗