Lipopolysaccharide inhibits proliferation of the cultured vascular smooth muscle cells by stimulating inducible nitric oxide synthase and subsequent activation of guanylate cyclase

This study was undertaken to investigate the mechanism of lipopolysaccharide (LPS) and nitric oxide (NO) as a regulator of vascular smooth muscle cell (VSMC) proliferation. VSMC was primarily cultured from rat aorta and confirmed by the immunocytochemistry with anti-smooth muscle myosin antibody. The number of viable VSMCs were counted, and lactate dehydrogenase (LDH) activity was measured to assess the degree of cell death. Concentrations of nitrite in the culture medium were measured as an indicator of NO production. LPS was introduced into the medium to induce the inducible nitric oxide synthase (iNOS) in VSMC, and Western blot for iNOS protein and RT-PCR for iNOS mRNA were performed to confirm the presence of iNOS. Inhibitors of iNOS and soluble guanylate cyclase (sGC), sodium nitroprusside (SNP) and L-arginine were employed to observe the action of LPS on the iNOS-NO-cGMP signalling pathway. LPS and SNP decreased number of VSMCs and increased the nitrite concentration in the culture medium, but there was no significant change in LDH activity. A cell permeable cGMP derivative, 8-Bromo-cGMP, decreased the number of VSMCs with no significant change in LDH activity. L-arginine, an NO substrate, alone tended to reduce cell count without affecting nitrite concentration or LDH level. Aminoguanidine, an iNOS specific inhibitor, inhibited LPS-induced reduction of cell numbers and reduced the nitrite concentration in the culture medium. LY 83583, a guanylate cyclase inhibitor, suppressed the inhibitory actions of LPS and SNP on VSMC proliferation. LPS increased amounts of iNOS protein and iNOS mRNA in a concentration-dependent manner. These results suggest that LPS inhibits the VSMC proliferation via production of NO by inducing iNOS gene expression. The cGMP which is produced by subsequent activation of guanylate cyclase would be a major mediator in the inhibitory action of iNOS-NO signalling on VSMC proliferation.

Nitroxergic nerve relaxes rat gastric smooth muscle by NO-cGMP pathway

This study was undertaken to investigate an involvement of nitroxergic innervation in gastric smooth muscle of rat. Isometric tension study, the measurement of single cell length, NADPH diaphorase stain of smooth muscle layers and neuronal nitric oxide synthase (nNOS) western blotting were performed. Sodium nitroprusside (SNP), a nitric oxide donor, relaxed the muscle strips precontracted by acetylcholine (ACh) in a concentration-dependent manner. Pretreatment of L-arginine decreased the contraction induced by electric field stimulation (EFS). Pretreatment of NG-nitro-L-arginine methyl ester (L-NAME), a NOS inhibitor, increased the EFS-induced contractions. LY 83583, a guanylate cyclase (GC) inhibitor, reversed the inhibitory actions of L-arginine on the muscle contractions. The effects of L-Arginine, L-NAME and LY 83583 on ACh-induced contractions were not significant. L-arginine reduced the EFS-induced contraction in circular muscle, whereas L-NAME enhanced the EFS-induced contraction in longitudinal strips. By EFS, the phasic contractions appeared approximately 20~25 seconds later. L-NAME significantly shortened the delay time to about 2~3 seconds. In single cell study, ACh contracted gastric smooth muscle cells, SNP relaxed the cells, and the latter also inhibited the ACh-induced contraction. LY 83583 enhanced the ACh-induced contraction and antagonized SNP-induced relaxation. NADPH diaphorase activity was assessed by a histochemistry, nitroblue tetrazolium (NTB) staining. Positive staining was observed in both circular and longitudinal muscle layers. L-arginine increased the staining, while L-NAME decreased the staining. Western blotting for nNOS proved the presence of nNOS in rat gastric smooth muscle. EFS and additional Ca2+ increased nNOS protein expression. These results suggest that in rat stomach, both circular and longitudinal muscle layers are innervated with nitroxergic nerves which relax the gastric smooth muscle via NO-cGMP pathway.

Effect of imipramine on calcium utilization of single cells isolated from canine detruso

This study is to investigate the mechanism of inhibitory effect of imipramine on the calcium utilization in single cells isolated from canine detrusor. 2 mm thick smooth muscle chops were incubated in 0.12% collagenase solution at 36degreeC, and aerated with 95% O2/5% CO2, and then cell suspension was examined Acetylcholine (ACh) evoked a concentration-dependent contraction of the isolated detrusor cells in normal physiologic salt solution (PSS), and the ACh-induced contraction was significantly inhibited by imipramine. In Ca2+-free PSS, ACh-induced contraction was less than those in normal PSS and it was not affected by the pretreatment with imipramine. Ca2+-induced contraction in Ca2+-free PSS was supressed by imipramine, but addition of A 23187, a calcium ionophore, overcomed the inhibitory effect of imipramine. High potassium-depolarization (40 mM KCl) evoked cell contraction, which was inhibited by imipramine. Caffeine, a releasing agent of the stored Ca2+ from sarcoplasmic reticulum, evoked a contraction of the cells that was not blocked by the pretreatment with imipramine. These results suggest that imipramine inhibits the influx of calcium in the detrusor cells through both the receptor-operated- and voltage-gated-calcium channels, but does not affect the release of calcium from intracellular storage site.

The Effects of Anticholinesterase Drugs on Gastric Motility / 영남의대학술지

BACKGROUND: Anticholinesterase drug inhibits acetylcholinesterase(AChE), induce accumulation of acetylcholine(ACh) near cholinergic receptors and cholinergic stimulation. This experiment was performed to study the effects of anticholinesterase drugs on gastric motility and the effect of ethanal on anticholinesterase drug-induced motility change. MATERIALS AND METHODS: After excision of stomach, 2x10mm circular musele strips were made, which were then fixed to the isolated muscle chamber. An isometric tension transducer was used to measure the contraction change of the gastric smooth muscle strips after drug addition. RESULTS: Fenthion, and irreversible anticholinesterase drug, increased ACh induced contraction of gastric smooth muscle strips and PAM, a cholinesterase activator, antagnized this action. Physostigmine, a reversible anticholinesterase drug, also increased the ACh induced contraction. The gastric motility was decreased by PAM. Ethanol, which is known to induce smooth muscle relaxation, inhibited the increase of contraction by fenthion. CONCLUSION: These results indicate that irreversible and reversible anticholinesterase drugs increase gastric motility and antagonized by cholinesterase activating drugs. And when exposed to both ethanol and anticholinesterase drug, gastric motility was decreased by the smooth muscle relaxation effect by ethanal.

The Effects of Ethanol on Cholinesterase Inactivation by Organophosphorous / 영남의대학술지

BACKGROUND: In korea the agricultural community widely uses organophosphorous, and organophosphorous poisonings are increasing every year. We compared change in activity of acetylcholinesterase and pseudocholinesterase by organophosphorous and by the interaction of ethanol and organophosphorous. We also compared the effect of reversible anticholinesterase drugs, physostigmine and neostigmine. The object of this study is to investigate the effects of several anticholinesterase drugs and on how ethanol influences the activity of cholinesterase. MATERIALS AND METHODS: Fifteen male university students were randomly selected, and blood samples were taken from the antecubital vein. The acetylcholinesterase in the RBC and the pseudocholinesterase in the serum were extracted and separated. The enzyme activity change was measured by the electrometric method. After adding acetylcholine, the pH change was measured with a pH meter. RESULTS AND CONCLUSION: Our results indicated that reversible anticholinesterase drugs decreased the cholinesterase activity more efficiently than organophosphorous. The acetyl cholinesterase and pseudocholinosterase activity were decreased by ethanol. When ethanol was added, oxime a cholinesterase activator, increased acetylcholinesterase activity but dose not increased pseudocholinesterase activity.

Modulatory Effect of Ammonium Carbonate on the GABAA Receptor

PURPOSE: This study was aimed to investigate the modulatory effect of ammonium carbonate on the GABAA receptor. METHODS: The effects of ammonium carbonate on the binding of radioligands to components of the GABAA receptor complex were observed. RESULTS: [3H]Flunitrazepam binding to the benzodiazepine receptor was enhanced by ammonium (1mM) ammonia concentrations. This increase in GABAergic neurotransmission is consistent with the clinical picture of lethargy, ataxia and cognitive deficits associated with liver failure and congenital hyperammonemia.

Effect of NG-nitro-L-arginine methyl ester and Methylene Blue on the Endotoxin-induced Vascular Hyporesponsiveness / 영남의대학술지

The study was undertaken to examine the intensity of involvement of inducible nitric oxide synthase(iNOS) and cyclic GMP signal transduction pathway as one of the mechanisms of vaso-relaxative action of bacterial lipopolysaccharide (LPS) on the canine femoral artery strips. Canine femoral arteries were isolated and spiral strips of 10 mm long and 2 mm wide were made in the Tyroad solution of 0-4degrees C. The strips were prepared for isometric myography in Biancani's isolated muscle chamber contaning 1 ml of Tyrode solution, which was maintained with pH 7.4 by areation with 95% O2/5% CO2 at 37degrees C and nitric oxide (NO) production was measured simulltaneously with isolated nitric oxide mrter. LPS induced NO production, suppressed the phenylephrine (PE) induced contraction and enhanced the acetylcholine (ACh) induced relaxation. NG-nitro-L-arginine methyl ester (L-NAME), an NOS inhibitor, methylene blue, a guanylyl cyclase inhibitor, potentiated PE induced contraction and suppressed ACh induced relaxation on the LPS treated strips. The inhibitory potency of methylene blue for LPS induced vascular hyporeponsiveness was stronger than that of L-NAME. These result suggest that in canine femoral artery, both iNOS and cyclic GMP signal transduction pathway are related with LPS indused vascular hyporeponsiveness, but in minor with iNOS and in major with cyclic GMP signal transduction pathway.

Changes of renal peripheral benzodiazepine receptor in the stress/anxiety response

Peripheral benzodiazepine receptor(PBR) has been identified in various peripheral tissues including kidney. The physiological and pharmacological functions of PBR are still uncertain, although it has been suggested that these are associated with the regulation of stress/anxiety response. Diazepam progeny, which were exposed to diazepam perinatally, was reported to be an animal model of chronic anxiety. However, PBR in the diazepam progenies are not known yet. In the present study, therefore, we examined the changes of PBR in the stress/anxiety response. Dams of rats were given injection of diazepam or vehicle during puerperium. Diazepam progenies showed increased level of anxiety on the performance of elevated plus maze, and increased Bmax of PBR. Saturation experiments followed by scatchard analysis of the results showed that the increase in the density of PBR and the affinity of the PBR remained unchanged. Forced swim stress increased anxiety on the plus maze in both groups of rats. In contrast to control, diazepam progenies did not show further upregulation of renal PBR immediately after swimming stress, but still higher than control. From the above results, it may be concluded that upregulation of renal PBR is associated with chronic anxiety as well as stress-induced response.

Effect of UV irradiation and rebamipide on the blood flow and viability of rabbit skin flap

This study was undertaken to examine the effects of ultraviolet light (UVL) and rebamipide on the cutaneous blood flow and tissue survival on rabbit skin flap. In a random bipedicle flap, Laser Doppler Flowmetry (LDF) was employed to measure the blood flow of flap (BFF). Wound Margin Strength (WMS) measured by force transducer and Light microscopy were used for evaluation of tissue viability. Single exposure to UVL increased the BFF gradually for more than 15 hours, and decreased the vasoconstrictor effect of intravenous phenylephrine. The UVL-induced increase in BFF regressed after 18 hours of irradiation, and this regression was tended to be enhanced by intradermal injection of L-NAME, a nitric oxide synthase (NOS) inhibitor, but the regression was significantly reversed by acetylcholine, an endothelial constitutive NOS (cNOS) activator and L-arginine, an NO precursor. Rebamipide, a novel antiulcer oxide synthase (NOS) inhibitor, but the regression was significantly reversed by acetylcholine, an endoagent known to scavenge the hydroxyl radical, abruptly reversed the spontaneous regression of the UVL-induced increase in BFF by the same manner as L-arginine. In ischemic skin flap, rebamipide increased the BFF abruptly by the same manner as sodium nitroprusside (SNP), an NO doner, while N-acetylcystein (NAC), a free radical scavenger, gradually increase the BFF. The rebamipide-induced increase in BFF was sustained at the level of the SNP-induced increase in BFF during the late period of experiment. Rebamipide increased the WMS of skin flaps and prevented the tissue necrosis in comparison with L-NAME. Based on these results, it is concluded that in rabbit skin, UVL irradiation increases the BFF by NO release, and rebamipide exerts a protective effect on the viability of ischemic skin flaps by either or both the increase in BFF by NO release and free radical scavenger effect.

Involvement of peripheral benzodiazepine receptor on contractility of canine trachealis muscle

Non-neuronal high affinity binding sites for benzodiazepines have been found in many peripheral tissues including cardiac muscle and vascular smooth muscle, and have been designated as 'peripheral benzodiazepine receptor'. Benzodiazepines have been shown to induce relaxation of the ileal, vesical, and uterine smooth muscles. However, it is still unclear about possible involvement of peripheral benzodiazepine receptor on the contractility of trachealis muscle. This study was performed to investigate the role of the peripheral benzodiazepine receptor on the contractility of canine trachealis muscle. Canine trachealis muscle strips of 15 mm long were suspended in an isolated organ bath containing 1 ml of physiological salt solution maintained at 37degreeC, and aerated with 95% O2/5% CO2. Isometric myography was performed, and the results of the experiments were as follows: Ro5-4684, FGIN-1-27 and clonazepam reduced a basal tone of isolated canine trachealis muscle strip concentration dependently, relaxant actions of Ro5-4684 and FGIN-1-27 were antagonized by PK11195, a peripheral benzodiazepine receptor antagonist. Flumazenil, a central type antagonist, did not antagonize the relaxant action of peripheral type agonists. Saturation binding assay of (3H)Ro5-4864 showed a high affinity (Kd = 5.33 +/- 1.27nM, Bmax = 867.3 +/- 147.2 fmol/mg protein) binding site on the canine trachealis muscle. Ro5-4684 suppressed the bethanechol-, 5-hydroxytryptamine- and histamine-induced contractions. Platelet activating factor (PAF) exerted strong and prolonged contraction in trachealis muscle strip. Strong tonic contraction by PAF was attenuated by Ro 5-4684, but not by WEB 2086, a PAF antagonist. Based on these results, it is concluded that the peripheral benzodiazepine receptor mediates the inhibitory regulation of contractility of canine trachealis muscle.

Effect of Baclofen on the Cholinergic Nerve Stimulation in Isolated Rat Detrusor / 영남의대학술지

This study aimed to investigate the mechanism of action of baclofen on the detrusor muscle isolated from rat. Rats (Sprague-Dawley) were sacrificed by decapitation and exsanguination. Horizontal muscle strips of 2 mm x 15mm were prepared for isometric myography in isolated muscle chamber bubbled with 95% / 5%-OZ / CO2 at 371C, and the pH was maintained at 7.4 Detrusor strips. contracted responding to the.. electrical field stimulation (EFS) by 2 Hz, 2U msec, monophasic square wave of 60 VDC. The initial peak of EFS-Induced contraction was tended to be suppresed by a,p-methylene-adenosine 5'-triphosphate (mATP), a partial agonist of purinergic receptor, and baclofen, a GABAB receptor agonist (statistically nonsignificant). The late sustained contraction by EFS was suppressed significantly (p < 0.05) by additions of atropione, a cholinergic muscarinic receptor antagonist and baclofen. The adenosine 5'-triphosphate-induced contraction was completely abolished by mA TP but not by baclofen. In the presence of atropine, the subsequent addition of acetylcholine could not contract the muscle strips: but the addition of acetylcholine in the presence of baclofen evoked a contraction to a remarkable extent.

Effect of Imipramine on the Contractility of Single Cells Isolated for Canine Detrusor / 영남의대학술지

The objective of this study was to establish a good methodology to isolate single smooth muscle cells that are alive and respond properly to pharmacological agents. Canine urinary bladders were employed as the source of single cells, and acetylcholine, atropine and imipramine were used as indicators of pharmacological responsiveness. Imipramine, an antidepressant drug exhibited the anticholinergic and calcium antagonizing properties on rat detrusor muscle. To establish a control value for a further experiment to elucidate the mechanism of action of imipramine on detrusor muscle, we measured the concentration-response of single cells to acetylcholine in the presesnce of imipramine by length of the cells and compared the result with the response in the presence of atropine. Tiny chops of smooth muscle taken from anesthetized canine urinary bladder were incubated in collagenase solution at 36℃ for 17-20 minutes. The collagenase solution included collagenase 1.2 mg/ml, soybean tryspin inhibitory 0.08 mg/ml, bovine serum albumin 2% in 10 ml Krebs-Henseleit buffer solution aerated with a consistent breeze of 95/5% O2/CO2 to maintain the pH at 7.4. After washing with plain K-H solution on 450 mesh, cells were dissociated from the digested tissue for 12-15 minutes. Cell suspension was transfered in 5 ml test tubes and acetylcholine was added for the final concentration to be 10⁻¹⁴~10⁻⁹M. To find the optimal time to fix the cells to determine the contractile responses, 1% acrolein was added 5, 10, 20, 30, 60 and 120 seconds after the administration of ACh. The length of cells fixed by acrolein were measured by microscaler vis CCTV camers on phaes-contrast microscope. The average length of 50 cells from a slide glass was taken as the value of a sample at the very concentration point. Single cells were isolated from canine detrusor. The length of untreated cells varied from 82 µm to 94 µm. The maximal response to actylcholine 10E-9M was accomplished within 5 seconds of exposure, and the shortening was 19±3%. Atropine reduced the contraction of the cells concentration-dependently. Lmipramine which exerts a cholinergic blocking action on some smooth muscles also reduced the contraction concentration-dependently and by a similar pattern as atropine. These findings document that imipramine may exerts a cholinergic blocking activity in the single smooth muscle cells isolated from canine urinary bladder.

Effect of GABA on the Contractility of Canine Trachealis Muscle / 영남의대학술지

This study aimed to investigate the existence of GABA receptor and the mechanisms of action of GABA and diazepam of the trachealis muscle isolated from dog. Horizontal muscle strips of 2mm×15mm were prepared from canine trachea, and isometric myography in isolated muscle chamber bubbled with 95/5%-O₂/CO₂ at 36℃, at the pH of 7.4 was performed. Muscle strips contracted responding to the electrical field stimulation (ESP) by 2~20 Hz, 20 msec, monophasic square wave of 60 VDC. GABA and diazepam suppressed the EFS-induced contractions to the similar extent, significantly. (p<0.05). Bicuculline, a GABA(A) receptor antagonist blocked both GABA- and diazepam-inhibitions; but DAVA, a GABA(B) receptor antagoinst did not affect either of them. These results suggest than in the canine trachealis muscle, there may be only GABA(A) receptor, and GABA and diazepam inhibit the contractility via GABA(A) receptor.

Characteristics of Potassium Channel in the Isolated Rat Detrusor Muscle / 영남의대학술지

The purpose of this study was to investigate the characteristics or the potassium channels existing in the rat urinary bladders. Smooth muscle strips of rat detrusor urinae were examined by isometric myography. Relaxation responses of detrusor muscle strips to the three potassium channel openers pinacidil, a cyanoguanidine derivative, BRL 38227, a benzopyran derivative and RP 52891, a tertrahydrothiopyran derivative were examined. The potassium channel openers reduced the basal tone, and the rank order of potency was RP 52891>pincidil>BRL 38227. Procaine, an inhibitor of the voltage-sensitive potassium channel tended to increase the basal tone, but it did not affect the relaxant effects of the calcium-activated potassium channel opener did not antagonize the relaxant effects, but it reduced the Emax of RP 52891 and BRL 38227. Glibenclamide, an inhibitor of the ATP-sensitive potassium channel, antagonized the relaxant effects of pinacidil, RP 52891 and BRL 38227 reducing the Emax of RP 52891 and BRl 38227. Galanin which inhibits secretion of insulin through opening the ATP-sensitive potassium channels in pancreatic β-cells rather increased the basal tone of the isolated detrusor strips. These results suggest that the urinary bladder of the rat has mainly the ATP-sensitive, glibenclamide sensitive potassium channel, which is a different type from that in the pancreatic β-islet cells.

Effects of octreotide on the contractility of isolated rat vas deferens / 영남의대학술지

This study was performed to investigate the effect of octreotide on the contractility of rat vas deferens. The -smooth muscle strips isolated from the prostatic portion were myographied in isolated organ bath. Electric -field stimulation (monophasic square wave, duration : 1. mSec, voltage : 50 V, frequency : 5 Hz or 30 Hz, train : 10 Sec) produced reproducible contraction. The contraction was composed of two component, first phasic component (FPC) and second tonicc component (STC).. These contractions were abolished by -tetrodotoxin (1 microM). Octreotide inhibited the field stimulation induced contractions both FPC and STC concentration- dependently. The FPC was decreased by a desentization of purinergic receptor by pretreatment of mATP, and the STC was decreased by pr,,creatment of reserpine (3 mg/kg, EP) 24 hours before experiments. Octreotide reduced the field stimulation induced contraction in the presence of mATP and of reserpinized muscle strips. The inhibitory effect of octreotide was more potent at 5 Hz than at 30 Hz. Octreotide did not affect basal ton and exogenous norepinephrine- or ATP-induced contraction. These results suggest that octreotide inhibit the contractility of the isolated rat vas deferens by inhibition of the release of neurotransmitters, both ATP and norepinephrine from adrenergic nerve terminal.

Effect of diazepam on the oxytocin induced contraction of the isolated rat uterus / 영남의대학술지

This study was designed to investigate the effect of diazepam on the spontaneous contraction and oxytocin induced contraction of the isolated rat uterus. Female rat (Sprague-Dawley) pretreated with oophorectomy and 4 days administration of estrogen. Weighing about 200 g, was sacrificed by cervical dislocation, and the uteruses were isolated. A longitudinal muscle strip was placed in temperature controlled (37℃) muscle chamber containing Locke's solution and myographied isometrically. Diazepam inhibited the spontaneous contraction and oxytocin induced contraction of the isolated rat uterus in a concentration-dependent manner. GABA, muscimol, a GABA A receptor agonist, bicuculline, a competitive GABA A receptor antagonist, picrotoxin, a non competitive GABA A receptor antagonist, baclofen, a GABA B receptor agonist, and delta-aminovaleric acid, a GABA B receptor antagonist, did not affect on the spontaneous and oxytocin induced contraction of the isolated rat uterus. The inhibitory actions of diazepam on the spontaneous and oxytocin induced contraction were not affected by all the GABA receptor agonists and antagonists, but exceptionally potentiated by bicuculline. This potentiation-effect by bicuculline was not antagonized by muscumol. In normal calcium PSS, addition of calcium restored the spontaneous contraction preinhibited by diazepam and recovered the contractile of oxtrocin preinhibited by diazepam. A23187, a calcium inophore, enhanced the restoration of both the spontaneous and oxytocin induced contraction by addition of calcium. In calcium-free PSS, diazepam suppressed the restoration of spontaneous motility by addition of calcium but allowed the recovery of spontaneous motility to a considerable extent. Diazepam could not inhibit some development of contractility by oxytocin in calcium-free PSS, but inhibited the increase in contractility by subsequent addition of calcium. These results suggest that the inhibitory action of diazepam on the rat uterine motility does not depend on or related to GABA receptors and that diazepam inhibits the extracellular calcium influx to suppress the spontaneous and oxytocin induced contractilities.

Inhibitory of γ-aminobutyric acid on the contractility of isolated rat vas deferens / 영남의대학술지

GABA is an inhibitory neurotransmitter in central nervous system and produce sedative, antianxiety and muscle relaxing effects via GABA(A) receptor or GABA(B) receptor. Recently it is known that GABA is widely distributed throughout peripheral organs and may play a physiological role in certain organ. The vas deferens is innervated by species-difference. These study, therefore, was performed to investigate the mode and the mechanism of action of GABA on the norepinephrine-, ATP- and electric stimulation-induced contraction of vas deferens of rat. Sprague-Dawley rats were sacrificed by cervical dislocation. The smooth muscle strips were isolated from the prostatic portion and were mounted in the isolated muscle bath. PSS in the bath was aerated with 95/5%-O₂/CO₂ at 33℃. Muscle tensions were measured by isometric tension transducer and were recorded by biological recording system. 1. GABA, muscimol, a GABA(A) agonist, and baclofen, a GABA(B) agonist inhibited the electric field stimulation (EFS, 0.2Hz, 1mSec, 80V, monophasic square wave)-induced contraction with a rank order of potency of GABA greater than baclofen greater than muscimol. 2. The inhibitory effect of GABA was antagonized by delta aminovaleric acid (DAVA), a GABA(B) antagonist, but not by bicuculline, a GABA(A) intagonist. 3. The inhibitory effect of baclofen was antagonized by DAVA, but the effect of muscimol was not antagonized by bicuculline. 4. Exogenous norepinephrine (NE) and ATP contracted muscle strip concentration dependently, but the effect of acetylcholine was negligible and GABA did not affect the NE-and ATP-induced contractions. 5. GABA, baclofen and muscimol did not affect basal tone, and GABA did not affect the NE-and ATP-induced contractions. 6. EFS-induced contraction was inclucling 2 distinctable components. The first phasic component was inhibited by beta gamma-methylene ATP (mATP), a desensitizing agent of APT receptor and the second tonic component was reduced by pretreatment of reserpine (3 mg/Kg, IP). 7. GABA inhibited the EFS-induced contraction of reserpinized strips, but not the mATP-treated strips. These results suggest that in the prostatic portion of the rat vas deferens, adrenergic and purinergic neurotransmissions are exist, and GABA inhibits the release of ATP via presynaptic GABA(B) receptor on the excitatory neurons.

Effect of GABA on the contratility of small intestine isolated from rat / 영남의대학술지

This study was designed to investigate the effect of GABA and related substances on the spontaneous contraction of rat small intestine. The rats (Sprague-Dawley), weighing 200-250g, were sacrificed by cervical dislocation, and the small intestine was isolated. Longitudinal muscle strips from duodenum, jejunum and ileum were suspended in Biancani's isolated muscle chambers and myographied isometrically. GABA and muscimol, a GABA A receptor agonist relaxed the duodenum and jejunum significantly, but baclofen-induced relaxation in those muscle strips negligible. The effectiveness of GABA and muscimol in various regions were the greatest on duodenum, and greater on jejunum than on ileum The effect of GABA and muscimol was antagonized by bicuculline, a competitive GABA A receptor antagonist and picrotoxin, a noncompetitive GABA A receptor antagonist. Duodenal relaxation induced by GABA and muscimol was unaffected by hexamethonium, but was prevented by tetrodotoxin. These results suggest that GABA inhibit the contractility of smooth muscle with distinct regional difference of efficacy, and the site of inhibitory action is the GABA A receptor existing at the presynaptic membrane of postganglionic excitatory nerves.

Existance of cholinergic and purinergic receptor on the detrusor muscle of rat urinary bladder / 영남의대학술지

This study was aimed at investigation of the stimulatory innervations on the rat urinary bladder. Detrusor muscle strips of 15 mm long were suspended in isolated muscle chambers containing 1 ml of PSS maintained at 37℃ and aerated with 95% O²/5% Co². Isometric myography was performed, and the results were as followings: Muscle strips showed “on-contraction” by electric field stimulation (EFS) frequency-dependently. The EFS-induced contraction was not affected by hexamethonium, a ganglion blocker, but abolished by tetrodotoxin, a nerve conduction blocker. Physostigmine, a cholinesterase inhibitor enhanced the EFS-induced contraction which was inhibited by hemicholinium, an inhibitor of choline uptake at the cholinergic nerve ending. Such an EFS-induced contraction was antagonized by atropine only partially, and the atropine-resistant portion was completely abolished by the desensitization of purinergic receptors by prolonged incubating of the strips in the presence of high concentration of ATP. Bethanechol, a cholinergic agonist, elicited concentration-dependent contraction. Adenosine triphosphate (ATP), a purinergic agonist, induced a weak but concentration-dependent contraction of short duration. Bethanechol-induced contraction was not affected by ATP-desensitization, and ATP-induced contraction was not affected by tetrodotoxin. These results suggest that there are at least two main stimulatory components of innervations in the detrusor muscle, cholinergic muscarinic and purinergic; and those receptors are independent each other.

The Effects of Diazepam on the Carbachol Induced Contraction of the Isolated Rat Ileum / 영남의대학술지

To investigate the effect of diazepam on the contractility of the intestinal smooth muscle, longitudinal muscle strip isolated from rat ileum was prepared for myography in isolated organ bath. 1) Basal tone of ileal muscle was reduced by diazepam concentration-dependently. 2) Higher concentrations (30 and 100 microM) of diazepam inhibited (p<0.05, p<0.001) The carbachol-induced contraction in a concentration-dependent manner; but lower concentration of diazepam (10 microM) enhanced (p<0.05). 3) Histamine-induced contraction was inhibited by pretreatment with diazepam in a concentration-dependent manner. 4) Ca⁺⁺-induced tension recovery in calcium-free solution was inhibited in the presence of diazepam concentration-dependently. These results suggest diazepam reduces the contractility of the longitudinal muscle isolated from rat ileum via interference with influx of calcium into the muscle cells.