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E E Daniel

Publications and source records attributed to E E Daniel.

At least 55 records · Page 3Linked to original sources
PubMed1998

Role of nitric oxide mechanisms in control of pyloric motility and transpyloric flow of liquids in conscious dogs.

The role of nitric oxide (NO) mechanisms in control of pyloric function and transpyloric flow were investigated in six conscious dogs. Antropyloroduodenal motility, transpyloric flow, and gastric emptying were measured 15 min after intravenous injection of 100 ml of either saline, L-arginine (50 mg/kg), L-NNA (5 mg/kg), or L-arginine (50 mg/kg) followed by L-NNA (5 mg/kg). Infusion of L-NNA was associated with retardation of gastric emptying (65 +/- 6%) in the first 30 min, in comparison to the saline (90 +/- 3%) or L-arginine (90 +/- 2%). This effect was prevented by infusion of L-arginine prior to L-NNA, after which 89 +/- 3% of the liquid emptied in 30 min. There was a significant reduction (P < 0.05) in the number and volume of flow pulses, and an increase in pyloric tone (P < 0.05) after L-NNA in comparison to the other three test conditions. There were no differences, however, in the number of antropyloric or isolated pyloric pressure waves under the four conditions. Our findings suggest that NO mechanisms influence gastric emptying and transpyloric flow of nonnutrient liquids by altering the pyloric tone, thus increasing resistance to flow.

Animals↗
PubMed1998

Myogenic nitric oxide synthase activity in canine lower oesophageal sphincter: morphological and functional evidence.

1. Studies on canine lower oesophageal sphincter (LOS) evaluated the existence and function of a myogenic, nitric oxide synthase (NOS) by use of immunocytochemistry for NOS isozymes, NADPH-d histochemistry, [3H]-L-arginine to [3H]-L-citrulline transformation. In addition, functional studies in the muscle bath were performed. 2. Smooth muscle bundles or freshly isolated smooth muscle cells of LOS were NADPH-d reactive but did not recognize some antibodies against neural, endothelial or inducible NOS. NADPH-d reactivity and immunoreactivity to a neural NOS antibody were colocalized in LOS enteric nerves. Muscle plasma membrane-enriched fractions from fresh and cultured LOS cells converted [3H]-L-arginine to [3H]-L-citrulline; activity was mostly Ca2+/calmodulin-dependent. 3. N-Nitro-L-arginine (L-NOARG) persistently increased tone (blocked by L-arginine) in muscle strips despite blockade of nerve function. Nifedipine prevented or abolished L-NOARG-induced, but not carbachol-induced, contraction showing that tone increase by L-NOARG required functional L-Ca channels. 4. Membrane-bound, myogenic NOS in canine LOS may release NO continuously when Ca2+ entry through L-Ca channels occurs under physiological conditions and thereby modulate tone in LOS.

Animals↗
PubMed1998

Myogenic NOS in canine lower esophageal sphincter: enzyme activation, substrate recycling, and product actions.

Depolarization elicited outward K+ currents from canine lower esophageal sphincter (LES) muscle cells, primarily through iberiotoxin (IbTX)- and tetraethylammonium-sensitive Ca(2+)-dependent K+ channels. Current magnitudes varied with pipette Ca2+ concentration (EC50 = 108.5 nM). NG-nitro-L-arginine (L-NNA, 10(-4)M), IbTX (10(-8)M), or buffering intracellular Ca2+ to 8 nM decreased outward currents > 80%. Sodium nitroprusside (NaNP, 10(-4)M) restored L-NNA-inhibited or low intracellular Ca2+ concentration (not IbTX)-inhibited currents. L-NNA or IbTX application depolarized LES cells from -43 to -35 mV. NaNP restored the membrane potential to -46 mV after L-NNA but not after IbTX application. Nifedipine (30 microM) reduced outward currents and abolished or reduced L-NNA or NaNP effects, respectively. Immunocytochemistry revealed the presence of both argininosuccinate synthetase and argininosuccinate lyase in LES muscle cells. L-Citrulline, like L-arginine, reversed L-NNA inhibition of outward currents; only L-arginine reversed inhibition of outward currents by an antibody to argininosuccinate synthetase. Therefore, endogenous nitric oxide production, activated by Ca2+ entrance involving L-type Ca2+ channels, may continuously enhance outward currents to modulate LES muscle cell membrane potential and excitability.

Animals↗
PubMed1998

Locations and molecular forms of PACAP and sites and characteristics of PACAP receptors in canine ileum.

In canine ileum we investigated the distribution of pituitary adenylate cyclase-activating peptide (PACAP), using immunofluorescence and radioimmunoassay and the binding of 125I-PACAP-27 to membranes. Nerve profiles immunoreactive to PACAP-27, and often to vasoactive intestinal polypeptide (VIP) as well, were found in all plexi, but PACAP was present in approximately 100-fold lesser amounts than VIP. High-performance liquid chromatography analysis of deep muscular plexus (DMP) synaptosomes suggested the presence of PACAP-38, PACAP-27, and a third unidentified molecular form. High- and low-affinity 125I-PACAP-27 binding sites were found in DMP synaptosomes and circular smooth muscle (CM) plasma membranes. In competition studies with DMP membranes, high (H)- and low (L)-affinity dissociation constants (Kd) and maximal binding capacities (Bmax) were as follows: KdH = 66.9 pM, BmaxH = 101 fmol/mg; KdL = 2.18 nM, BmaxL = 580 fmol/mg protein. The binding of 125I-PACAP-27 was fast. Dissociation was slow and incomplete in the presence of unlabeled PACAP-27 but accelerated by pretreatment with guanosine 5'-O-(3-thiotriphosphate) (GTP gamma S). GTP gamma S or cholera toxin treatment eliminated high-affinity binding in both membranes. VIP had approximately 100-fold lower affinity than PACAP-27 in both membranes. Cross-linking studies identified an approximately 70-kDa PACAP receptor in each membrane. Thus PACAP coexists with VIP in ileal enteric nerves and acts on PACAP-preferring, possibly Gs-coupled, receptors in DMP synaptosomes and CM membranes.

Animals↗
PubMed1998

Role of gap junctions in structural arrangements of interstitial cells of Cajal and canine ileal smooth muscle.

We examined the structural and functional basis for pacemaking by interstitial cells of Cajal (ICC) in circular smooth muscle of the canine ileum. Gap junctions were found between ICC of myenteric plexus (MyP), occasionally between MyP ICC and outer circular smooth muscle cells, between individual outer circular smooth muscle cells, between them and ICC of the deep muscular plexus (DMP), and between DMP ICC. No visible gap junctions connected MyP ICC to longitudinal muscle cells or inner circular muscle cells. Occasionally contacts occurred between the two muscle layers. No special structures were found to connect MyP and DMP ICC networks. Octanol concentration dependently reduced the amplitude and frequency of, but did not abolish, slow waves in circular muscle in isolated ileum recorded near the MyP or the DMP. Slow waves triggered from MyP ICC by a current pulse also persisted. Contractile activity was abolished, cells were depolarized, and fast inhibitory junction potentials were reduced by octanol. We conclude that ICC pacemakers of the MyP and DMP utilize gap junctional conductances for pacemaking function but may not require them. Coupling between the two ICC networks may utilize the circular muscle syncytium.

Animals↗
PubMed1998

Release of epithelium-derived PGE2 from canine trachea after antigen inhalation.

To investigate the role of prostaglandin (PG) E2 in allergen-induced hyperresponsiveness, dogs inhaled either the allergen Ascaris suum or vehicle (Sham). Twenty-four hours after inhalation, some animals exposed to allergen demonstrated an increased responsiveness to acetylcholine challenge in vivo (Hyp-Resp), whereas others did not (Non-Resp). Strips of tracheal smooth muscle, either epithelium intact or epithelium denuded, were suspended on stimulating electrodes, and a concentration-response curve to carbachol (10(-9) to 10(-5) M) was generated. Tissues received electrical field stimulation, and organ bath fluid was collected to determine PGE2 content. With the epithelium present, all three groups contracted similarly to 10(-5) M carbachol, whereas epithelium-denuded tissues from animals that inhaled allergen contracted more than tissues from Sham dogs. In response to electrical field stimulation, Hyp-Resp tissues contracted less than Sham tissues in the presence of epithelium and more than Sham tissues in the absence of epithelium. PGE2 release in the muscle bath was greater in Non-Resp tissues than in Sham or Hyp-Resp tissues when the epithelium was present. Removal of the epithelium greatly inhibited PGE2 release. We conclude that tracheal smooth muscle is hyperresponsive in vitro after in vivo allergen exposure only when the modulatory effect of the epithelium, largely through PGE2 release, is removed.

Administration, Inhalation↗
PubMed1998

Pharmacological and immunocytochemical characterization of subtypes of alpha-1 adrenoceptors in dog aorta.

In this study, the effects of nine alpha-1 adrenoceptor antagonists [prazosin, WB 4101 (WB), chloroethylclonidine (CEC), 5-methylurapidil (5-MU), BMY 7378 (BMY), MDL 73005EF (MDL73), MDL 72832 (MDL72), RS 17053 (RS) and SK&F 105854 (SKF)] were studied on contractile responses to phenylephrine (PE) of the endothelium-denuded dog aorta in vitro. All antagonists, except CEC, 5-MU and RS, produced concentration-dependent competitive inhibition of contractile responses of the aorta to PE. The rightward shift of the concentration-response curves of PE yielded constant pKB values with increasing antagonist concentrations in most cases allowing a single pooled value to be determined: for prazosin, a pKB of 8.99 +/- 0.11 (n = 20, KB of 1.03 nM); for WB, a pKB of 8.75 +/- 0.08 (n = 23, KB of 1.76 nM); for BMY, a pKB of 7.21 +/- 0.13 (n = 13, KB of 62 nM); for MDL72, a pKB of 7.95 +/- 0.15 (n = 12, KB of 11.2 nM); and for SK&F 105854, a pKB of 5.82 +/- 0.08 (n = 15, KB of 1.52 microM). For MDL73, pKB values decreased with antagonist concentration: 7.88 +/- 0.06 at 10 nM, 7.56 +/- 0.28 at 100 nM and 6.92 +/- 0.18 at 1000 nM, which suggests the presence of more than one receptor subtype. CEC (10 and 100 microM) almost completely inhibited responses to PE; lower concentrations had no significant effect. 5-MU (10-300 nM) and RS (3-300 nM) were ineffective antagonists in this tissue. Because WB, a highly selective alpha-1D and alpha-1A adrenoceptor subtypes inhibitor, blocked PE responses (with less affinity than for alpha-1A adrenoceptors), and 5-MU and RS, which are selective blockers for alpha-1A adrenoceptor, were ineffective, we conclude that alpha-1A adrenoceptors are absent in the dog aorta. The effects of the less selective MDL72 were inconsistent with actions at alpha-1B or alpha-1D adrenoceptors. Although WB shifted the PE concentration-response curve to the right, the abilities of BMY, MDL73 and SKF to inhibit competitively PE contraction were of lower affinity compared with expectations for interaction with alpha-1D adrenoceptors; they are not the predominant subtype. The complete inhibition of PE responses by CEC suggests that the dog aorta contains the alpha-1B adrenoceptor subtype. In immunocytochemical studies of the expression of alpha-1B adrenoceptor, all cells apparently expressed this protein. Moreover, Western blot studies of the microsomal fractions confirmed the presence of alpha-1B adrenoceptors. In the dog aorta, the alpha-1 adrenoceptors predominantly resemble alpha-1B rather than alpha-1D adrenoceptors as reported in the rat aorta.

Animals↗
PubMed1998

Calcium source diversity in canine lower esophageal sphincter muscle.

Tonic contraction of the lower esophageal sphincter (LES) prevents gastroesophageal reflux. LES tone is produced both by cholinergic nerve and myogenic activities. The Ca++ sources for LES tone and carbachol-induced contraction in canine LES strips were determined from the effect on contractile activity of extracellular Ca++ level modulation, Ca++ entrance blockade or enhancement with nifedipine or BayK8644 respectively, and/or inhibition of Ca++ store refilling using the sarcoplasmic reticulum Ca++ pump inhibitor, cyclopiazonic acid. LES tone disappeared when a Ca++-free physiological saline solution or nifedipine was applied. Sustained Ca++ free contractions to carbachol were prevented/abolished by nifedipine or increased Ca++ chelation and enhanced by BayK8644. Inhibition of sarcoplasmic reticulum Ca++ pumps by cyclopiazonic acid reduced Ca++ free contractions to carbachol; BayK8644 restored cyclopiazonic acid-reduced Ca++ free contractions to carbachol. Therefore, some Ca++ stores can be refilled by mechanisms not requiring activity of the sarcoplasmic reticulum Ca++ pump. A preferred pathway may exist whereby Ca++ enters stores directly through L-Ca++ channels. The proposed Ca++ store refilling mechanism involves continuous Ca++ entry through L-Ca++ channels from sites not equilibrated with external Ca++. Therefore, diverse Ca++ stores exist in canine LES which are dependent on Ca++ influx through L-Ca++ channels.

3-Pyridinecarboxylic acid, 1,4-dihydro-2,6-dimethy↗
PubMed1997

Mobilization of internal Ca2+ by vasoactive intestinal polypeptide in endothelial cells.

The aims of the present study were to establish whether vasoactive intestinal polypeptide (VIP) could mobilize internally-stored Ca2+ and whether Ca2+ release could trigger Ca2+ influx from the extracellular space. Bovine pulmonary artery endothelial cells from an established cell line were loaded with fura-2/AM and cells were studied in suspension or were imaged in monolayers at 40-80% confluency. In Ca2+ imaging studies, VIP evoked Ca2+ transients in Ca2+-free medium containing 50 microM EGTA. This was observed in 33 out of 122 cells examined on 29 separate trials. With each cell, the spread of Ca2+ appeared to occur from the periphery of the cell to the central core. Cells which did not respond to VIP responded to other stimulants such as bradykinin, endoplasmic reticulum Ca2+ pump inhibitors, (cyclopiazonic acid and thapsigargin), and endoplasmic reticulum Ca2+ release channel opener, ryanodine. The reintroduction of Ca2+ following VIP-induced Ca2+ release did not evoke a Ca2+ response in 5 cells imaged. Cells in suspension showed typical biphasic responses to bradykinin, thapsigargin or cyclopiazonic acid in the presence of external Ca2+. Stimulation with VIP caused transient Ca2+ responses in Ca2+-free physiological saline containing 50 microM EGTA. However, only 1 out of 4 cells tested showed a response to Ca2+ when it was reintroduced to the bathing medium. This study provided direct evidence for the first time in these bovine endothelial cells for VIP-mediated elevation of cytosolic concentration of Ca2+. The results also suggested that other mechanisms might prevail preventing capacitative Ca2+ entry following the release of internally-stored Ca2+.

Animals↗
PubMed1997

The distribution of NPY-containing nerves and the catecholamine contents of canine enteric nerve plexuses.

Our previous study demonstrated that PYY was a major neuropeptide in the canine enteric nervous system, the present study defines the locations of NPY-containing enteric neurons. NPY-positive nerve cell bodies and fibers were numerous in gastric and pyloric myenteric plexuses as were positive nerve fibers in antral and pyloric muscle layers, pyloric sphincter muscle layers, and surrounding blood vessels. In contrast to findings for PYY, there were considerably fewer NPY-positive nerve cell bodies and fibers in the canine ileum and colon. Noradrenaline was the predominant catecholamine in all plexuses, the rank order of its contents being: deep muscular > submucous > myenteric plexus. The dopamine/noradrenaline ratio was constant in all plexuses; adrenaline was present in minor amounts. PYY is the more abundant neuropeptide in ileum and colon, and NPY has a greater presence in the gastric antrum, pylorus, and surrounding blood vessels. PYY and NPY may play different functional roles in the GI tract.

Animals↗
PubMed1997

Actions of 4-chloro-3-ethyl phenol on internal Ca2+ stores in vascular smooth muscle and endothelial cells.

1. Recently, 4-chloro-3-ethyl phenol (CEP) has been shown to cause the release of internally stored Ca2+ apparently through ryanodine-sensitive Ca2+ channels, in fractionated skeletal muscle terminal cisternae and in a variety of non-excitable cell types. Its action on smooth muscle is unknown. In this study, we characterized the actions of CEP on vascular contraction in endothelium-denuded dog mesenteric artery. We also determined its ability to release Ca2+, by use of Ca2+ imaging techniques, on dog isolated mesenteric artery smooth muscle cells and on bovine cultured pulmonary artery endothelial cells. 2. After phenylephrine-(PE, 10 microM) sensitive Ca2+ stores were depleted by maximal PE stimulation in Ca2+-free medium, the action of CEP on refilling of the emptied PE stores was tested, by first pre-incubating the endothelium-denuded artery in CEP for 15 min before Ca2+ was restored for a 30 min refilling period. At the end of this period, Ca2+ and CEP were removed, and the arterial ring was tested again with PE to assess the degree of refilling of the internal Ca2+ store. 3. In a concentration-dependent manner (30, 100 and 300 microM), CEP significantly reduced the size of the post-refilling PE contraction (49.4, 28.9 and 5.7% of control, respectively) in Ca2+-free media. This suggests that Ca2+ levels are reduced in the internal stores by CEP treatment. CEP alone did not cause any contraction either in Ca2+-containing or Ca2+-free Krebs solution. 4. Restoring Ca2+ in the presence of PE caused a large contraction, which reflects PE-induced influx of extracellular Ca2+. The contraction of tissues pretreated with 300 microM CEP was significantly less compared with controls. However, tissues pretreated with 30 and 100 microM CEP were unaffected. Washout of CEP over 30 min produced complete recovery of responses to PE in Ca2+-free and Ca2+-containing medium suggesting a rapid reversal of CEP effects. 5. Concentration-response curves were constructed for PE, 5-hydroxytryptamine (5-HT) and K+ in the absence of and after 30 min pre-incubation with 30, 100 and 300 microM CEP. In all cases, CEP caused a concentration-dependent depression of the maximum response to PE (84.8, 43.4 and 11.6% of control), 5-HT (65.4, 25.7 and 6.9% of control) and K+ (77.6, 41.1 and 10.8% of control). 6. Some arterial rings were pre-incubated with ryanodine (30 microM) for 30 min before the construction of PE concentration-response curves. In Ca2+-free Krebs solution, ryanodine alone did not cause any contraction. However, 58% (11 out of 19) of the tissues tested with ryanodine developed contraction (6.9+/-1.2% of 100 mM K+ contraction, n=11) in the presence of external Ca2+. EC50 values for PE in ryanodine-treated tissues (1.7+/-0.25 microM, n=16) were not significantly different from controls (2.5+/-0.41 microM, n=22). Maximum contractions to PE (118.5+/-4.4% of 100 mM K+ contraction, n=16) were also unaffected by ryanodine when compared to controls (129+/-4.2%, n=23). 7. When fura-2 loaded smooth muscle cells (n=13) and endothelial cells (n=27) were imaged for Ca2+ distribution, it was observed that 100 and 300 microM CEP in Ca2+-free medium caused Ca2+ release in both cell types. Smooth muscle cells showed a small decrease in cell length. Addition of EGTA (5 mM) reversed the effect of CEP on intracellular Ca2+ to control values. 8. These data show, for the first time in vascular smooth muscle and endothelial cells, that CEP releases Ca2+ more rapidly than ryanodine. Unlike ryanodine, CEP caused no basal contraction but depressed contractions to PE, 5-HT and K+. The lack of basal contraction may result from altered responsiveness of the contractile system to intracellular Ca2+ elevation.

Animals↗
PubMed1997

Characterization of alpha-adrenoceptors in canine mesenteric vein.

The alpha-adrenergic receptors (alpha-ARs) in canine mesenteric vein (DMV) were studied by using nonselective agonists and selective antagonists in functional studies and in ligand binding to classify the subtypes present. Based on functional studies of phenylephrine (PE)-induced contractions and ligand-binding interactions of [3H]-prazosin with prazosin (PR), WB 4101 (WB), 5-methylurapidil (5-MU), BMY 7378, and SK&F 105854, and pretreatment with chloroethylclonidine (CEC), DMV alpha1-ARs resembled the alpha1D subtype. However, the affinity of PR assessed in functional and ligand-binding studies was less (pK(B,D,i) < or = 9) than expected from previous characterization of cloned rat or human alpha1-AR (pKi > or = 10). Interactions with 5-MU, BMY 7378, or SK&F 105854 suggested the presence of some alpha1-ARs that were not typical of alpha1D-AR and that binding and functional interactions did not yield corresponding results. PR binding was abolished by treatment with CEC, contractile responses to PE were reduced in Emax, and the concentration-effect curve shifted to the left, as previously reported. DMVs contracted in response to alpha2-AR agonists and were studied when contractions were potentiated by increasing extracellular KCl to 20 mM. Rauwolscine (RAU) had K(B) values at these sites consistent with K(D) values in binding studies. CEC had no effect on RAU binding in DMV. Ligand-binding studies to [3H]-RAU sites did not reveal a clear identification of subtype, but these alpha2-ARs were clearly not alpha2B-ARs. We conclude that canine mesenteric vein contains alpha1D-like ARs, but with significant differences, and an unclassifiable alpha2-AR. There may also be a smaller population of other, not alpha1D-like ARs, receptors, mediating responses to PE and binding of prazosin.

Adrenergic alpha-Agonists↗
PubMed1997

Myogenic and neurogenic mechanisms and arachidonate metabolites in bronchial muscle response to allergen.

We investigated allergen-induced airway hyperresponsiveness (AH) in bronchial tissues obtained from dogs that inhaled Ascaris suum leading to AH (RESP) in vivo or that exhibited no change (NON-RESP) as well as from dogs that inhaled saline (SHAM). RESP tissues were not hyperresponsive to KCl or to carbachol, whereas contractions to electrical field stimulation (EFS) were reduced. This reduction was reversed partially by indomethacin and completely by replacement of the bathing fluid. Radioimmunoassay revealed marked elevation of prostaglandin (PG) E2 generation in RESP tissues compared with SHAM and NON-RESP tissues. EFS-evoked contractions were often followed by a slowly developing secondary contraction in RESP tissues but not in SHAM or NON-RESP tissues. However, indomethacin unmasked such secondary contractions in many SHAM and NON-RESP tissues and markedly enhanced those in RESP tissues, whereas L-655,240 (thromboxane A2/PGD2 receptor antagonist) abolished such contractions in all groups. We were unable to detect thromboxane using radioimmunoassay. We conclude that allergen-induced AH involves altered generation of cyclooxygenase metabolites of arachidonic acid (particularly PGE2) as well as of a nonprostanoid inhibitory factor; as such, the responsiveness of the tissue in vitro is dependent on the relative levels of inhibitory and excitatory metabolites.

Allergens↗
PubMed1997

Influence of nitric oxide and vasoactive intestinal peptide on the spontaneous and triggered electrical and mechanical activities of the canine ileum.

Modulation of canine ileal pacemaker activity by nitric oxide (NO) or vasoactive intestinal peptide (VIP) was studied during recording of the intracellular electrical and mechanical activity from the entire muscularis externa and from an isolated circular muscle preparation both cut in the long axis of the circular muscle. In the whole-thickness preparation with cholinergic and adrenergic nerve function blocked, the inhibitory junction potentials (IJPs) recorded near the myenteric plexus (MyP) or deep muscular plexus (DMP) were abolished by omega-conotoxin GVIA (omega-CTX, 10(-7) to 3 x 10(-7) M), tetrodotoxin (TTX, 1 microM), or the NO synthase (NOS) inhibitor N omega-nitro-L-arginine (L-NNA at 50 microM). IJPs from electrical field stimulation triggered slow waves (TSWs); after TTX or omega-CTX, TSWs still occurred, advanced in time and increased in amplitude after TTX. Addition of L-NNA advanced the onset of the TSWs after omega-CTX. TTX, L-NNA, or omega-CTX left the resting membrane potentials, the characteristics of spontaneous slow waves, or TSWs evoked by a long stimulating pulse unchanged. L-NNA at 100 microM enhanced the amplitude but not the frequency of spontaneous slow waves. TTX and NOS blockers all increased circular muscle contractions associated with the spontaneous slow waves and TSWs. In isolated circular muscle preparations, the NOS inhibitors N omega-nitro-L-arginine methyl ester (L-NAME at 300 microM) or L-NNA at 100 microM abolished the IJPs and increased the regularity and amplitude of spontaneous slow waves and associated contractions, but TSWs could not be evoked before or after NOS inhibition. The NO donor 3-morpholinosydnonimine hydrochloride (SIN-1) at 200 microM caused hyperpolarizations (10-15 mV) similar to the IJP mediator, attenuated the IJPs, and abolished mechanical activities. SIN-1 increased the slow wave frequency but decreased the amplitude and duration of spontaneous slow waves and TSWs. VIP (10(-6) M) decreased contraction and slow wave amplitude and prolonged IJP duration without affecting membrane potential or slow wave frequency. We conclude that spontaneous slow waves and TSWs originate independently of neural activity. Pacemaking regions possess inhibitory neural inputs that release NO to mediate IJPs and relaxation and influence the delay before a TSW. NO (not VIP) release from nerves inhibits initiation of spontaneous slow waves or TSWs near the MyP, and spontaneous NO release modulates pacemaking activity from the DMP.

Animals↗
PubMed1997

Sites of nitric oxide (NO) actions in control of circular muscle motility of the perfused isolated canine ileum.

In the isolated intra-arterially perfused canine ileum, N omega -nitro-L-arginine (L-NNA, 3 x 10(-4) M) increased tonic and phasic motor activity of the circular muscle. As has previously been shown, L-NNA enhanced contractions to electrical field stimulation at sites proximal to the serosal electrodes and converted initial relaxation when present at distal sites to contraction. L-NNA shifted the acetylcholine dose-response curve to the left and amplified the response to low-dose acetylcholine. Following L-NNA, addition of 10(-5) M sodium nitroprusside (NO donor) returned the tonic and phasic activity, the electrical field stimulation responses, and the acetylcholine dose-response curve to control values. Tetrodotoxin (TTX, 10(-6) M) increased tone (less than L-NNA) and abolished responses to both electrical field stimulation and motor activity induced by prior L-NNA. Subsequent L-NNA did not alter TTX-induced tonic motor responses. TTX also shifted the acetylcholine dose-response curve leftward and increased the responses to low-dose acetylcholine. After TTX, sodium nitroprusside returned the low-dose acetylcholine responses to control values and, after L-NNA, failed to restore them to control values. After L-NNA and TTX, sodium nitroprusside restored responses to low-dose acetylcholine to control values, Thus, removal of inhibition of the release of excitatory neurotransmitters, not removal of actions of NO on the muscle, accounted for the increases in tonic and phasic activity from L-NNA. Uninhibited release of excitatory transmitters augmented circular muscle responses to low-dose acetylcholine. TTX eliminated effects of excitatory transmitters, allowing exogenous NO to reduce low-dose acetylcholine contractions. No treatment affected the maximum responses to acetylcholine, produced by a contractile mechanism independent of muscle excitability and unaffected by exogenous NO or release of neurotransmitters.

Animals↗
PubMed1996

Plant alkaloids, tetrandrine and hernandezine, inhibit calcium-depletion stimulated calcium entry in human and bovine endothelial cells.

Depletion of internal Ca2+ stores causes capacitative Ca2+ entry which occurs through non-selective cation channels sensitive to blockade by SK&F 96365. Recently, alkaloids of Chinese herbal medicinal origin, tetrandrine and hernandezine, have been shown to possess actions including inhibition of Ca2+ channels in non-excitable cell types. In this study, we compared the actions of these novel inhibitors to those of SK&F 96365 in fura-2-loaded endothelial cells from human umbilical vein and bovine pulmonary artery. Depletion of Ca2+ from the internal stores was accomplished in Ca(2+)-free medium using an endoplasmic reticulum Ca2+ pump inhibitor, cyclopiazonic acid (CPA) or receptor agonists, histamine and bradykinin. Stimulation with histamine or bradykinin caused a marked and rapid transient increase in Ca2+ signal whereas CPA caused a smaller amplitude increase of longer duration. Restoring Ca2+ to the medium caused marked and sustained increases in the fluorescence indicating movement of Ca2+ into the cytosol presumably stimulated by the emptied Ca2+ stores. SK&F 96365 as well as tetrandrine and hernandezine antagonized depletion-induced Ca2+ entry. The results suggest that these putative inhibitors interact with Ca2+ entry triggered by depletion of the internal Ca2+ stores and their action is presumed to be on the non-selective cation channels. Their effectiveness may be enhanced by the mechanisms which lead to the opening of the Ca2+ influx channel.

Alkaloids↗
PubMed1996

Heterogeneity of motilin receptors in the gastrointestinal tract of the rabbit.

Motilin, a 22-amino acid peptide synthesized in endocrine cells of intestinal mucosa, stimulates GI smooth muscle contractility. To elucidate the mode of action of motilin, we attempted to determine whether motilin receptors are localized on nerve cells or on smooth muscle cells of the GI tract. Mucosa-free tissues from rabbit antrum and duodenum were homogenized separately with a Polytron prior to differential centrifugation to obtain synaptosome or plasma membrane-enriched fractions, as determined by the distribution of [3H]saxitoxin (SAX) binding (neural membranes) and 5' nucleotidase (5'N) activity (smooth muscle plasma membranes). Motilin binding was evaluated by the displacement of [125I]motilin by motilin (1-22) on the various membrane fractions. In the antrum, motilin binding was highly correlated with SAX binding (r = 0.81, p < 0.0005), and also significantly with 5'N activity (r = 0.54, p < 0.05). In the duodenum, motilin binding correlated significantly with 5'N activity (r = 0.67, p < 0.005), but not with SAX binding (r = -0.11, NS). Receptor affinity, for the motilin antagonist MOT(1-12)[CH2NH]10-11, for motilin(1-22), and for the motilin agonist erythromycin lactobionate was significantly (p < 0.001, p < 0.001, and p < 0.05, respectively) higher in SAX-enriched fractions from the antrum than in 5'N-enriched fractions from the duodenum. Therefore, in the rabbit: 1) motilin receptors appear to be predominantly located on nerve tissues in the antrum and restricted to smooth muscle cells in the duodenum, and 2) antral receptors and duodenal receptors displayed different pharmacological characteristics, probably corresponding to two specific and heterogeneous motilin receptor subtypes.

5'-Nucleotidase↗