vasoactive-intestinal-peptide and alpha-beta-methyleneadenosine-5--triphosphate

In the rat gastric fundus, non-adrenergic, non-cholinergic (NANC) relaxations are mediated by nitric oxide (NO), vasoactive intestinal polypeptide (VIP), and a third, as yet unidentified, neurotransmitter. The possible involvement of adenosine 5'-triphosphate (ATP) in the NANC relaxations was examined using pyridoxalphosphate-6-azophenyl-2',5'-disulphonic acid (PPADS), apamin and desensitization to alpha,beta-methylene ATP. NANC responses were studied in the absence and presence of N(G)-nitro-L-arginine methyl ester (NAME; 100 microM) and alpha-chymotrypsin (1 u ml(-1)), to inhibit responses to NO and VIP, respectively. PPADS (100 microM), apamin (1 microM) and desensitization to alpha,beta-methylene ATP (10 microM, three additions) all significantly (P<0.05) reduced NANC relaxations to electrical field stimulation (0.5 - 4 Hz, 30 s trains) in longitudinal strips of rat gastric fundus and almost abolished the residual relaxation remaining in the presence of NAME and alpha-chymotrypsin. PPADS had no effect on responses to the NO-donor, sodium nitroprusside (SNP), or VIP. Apamin slightly reduced relaxations to SNP, but did not affect those to VIP, whereas desensitization to alpha,beta-methylene ATP markedly reduced responses to both SNP and VIP. The effects of PPADS and apamin in this study provide strong evidence that the third inhibitory NANC neurotransmitter in the rat gastric fundus is ATP.

Topics: Adenosine Triphosphate; Animals; Apamin; Atropine; Dose-Response Relationship, Drug; Drug Interactions; Electric Stimulation; Gastric Fundus; Guanethidine; In Vitro Techniques; Male; Muscle Relaxation; Nerve Tissue; Neurotransmitter Agents; Nitroprusside; Platelet Aggregation Inhibitors; Pyridoxal Phosphate; Rats; Rats, Inbred SHR; Vasoactive Intestinal Peptide; Vasodilator Agents

The role of nitric oxide (NO) and ATP in the regulation of nonadrenergic, noncholinergic (NANC) inhibitory transmission in the pylorus remains unclear. In the presence of atropine and guanethidine, electric field stimulation induced NANC relaxations in a frequency-dependent manner (1-20 Hz) in the rat pylorus. NANC relaxations were significantly inhibited by N(G)-nitro-L-arginine methyl ester (L-NAME; 10(-4) M). P(2X) purinoceptor antagonist pyridoxal phosphate-6-azophenyl-2',4'-disulfonic acid (PPADS; 3 x 10(-5) M) and P(2Y) purinoceptor antagonist reactive blue 2 (2 x 10(-5) M) had no effect on NANC relaxations. However, the combined administration of L-NAME and PPADS, but not reactive blue 2, evoked greater inhibitory effects on NANC relaxation than that evoked by L-NAME alone. alpha-Chymotrypsin and vasoactive intestinal polypeptide antagonist did not affect NANC relaxations. ATP (10(-5)-10(-3) M) and P(2X) purinoceptor agonist alpha, beta-methyleneadenosine 5'-triphosphate (10(-7)-10(-5) M), but not P(2Y) purinoceptor agonist 2-methylthioadenosine 5'-triphosphate (10(-7)-10(-5) M), induced muscle relaxations in a dose-dependent manner, and relaxations were significantly reduced by PPADS and unaffected by TTX. These studies suggest that NO and ATP act in concert to mediate NANC relaxation of the rat pylorus. ATP-induced relaxation appears to be mediated by P(2X) purinoceptors located on smooth muscle cells.

Topics: Adenosine Triphosphate; Animals; Carbachol; Electric Stimulation; Male; Muscle Contraction; Muscle, Smooth; Neuropeptides; NG-Nitroarginine Methyl Ester; Nitric Oxide; Nitroprusside; Pituitary Adenylate Cyclase-Activating Polypeptide; Pylorus; Pyridoxal Phosphate; Rats; Rats, Sprague-Dawley; Receptors, Purinergic P2; Stomach; Tetrodotoxin; Thionucleotides; Vasoactive Intestinal Peptide

1. Purinergic and cholinergic components of parasympathetic neurotransmission and contractile responses to exogenous alpha,beta-methylene ATP, acetylcholine, substance K, substance P, calcitonin gene-related peptide, vasoactive intestinal polypeptide and capsaicin have been investigated in the urinary bladder of hibernating hamsters (4 weeks), cold exposed (4 weeks) and age-matched controls. 2. Electrical field stimulation (EFS) evoked increased frequency-dependent contractions in the detrusor strips from hibernating hamsters compared with those obtained from cold-exposed and age-matched animals. Tetrodotoxin (10(-6) M) completely blocked the frequency-dependent contractions in all groups. 3. The purinergic component of the parasympathetic neurotransmission was not affected in hibernating and cold-exposed animals while the cholinergic component was increased with respect to age-matched animals. The neurogenic response to EFS, still present after incubation with atropine (10(-6) M) and suramin (10(-4) M), was attenuated by indomethacin (10(-6) M) and blocked by tetrodotoxin (10(-6) M). 4. Exogenous administration of alpha,beta-methylene ATP elicited a significantly reduced contraction in strips from hibernating and cold-exposed hamsters relative to age-matched animals. The contractile response to exogenous acetylcholine was greater in the detrusors from hibernating hamsters than in cold-exposed and age-matched animals. Substance K elicited reduced contractions in preparations from hibernating animals compared with cold-exposed and control animals. Calcitonin gene-related peptide, vasoactive intestinal polypeptide, substance P and capsaicin did not elicit any relaxant or contractile response either at resting tone or in carbachol (5 x 10(-7) M)-precontracted tissues. 5. In summary, our findings indicate that 4 weeks of hibernation can significantly increase neurogenic responses in the hamster urinary bladder. This appears to be due to an increase in postjunctional responses to acetylcholine. In contrast, there was a decrease of the postjunctional responses to the parasympathetic cotransmitter ATP and also to the sensory-motor neurotransmitter substance K.

Topics: Acetylcholine; Adenosine Triphosphate; Animals; Calcitonin Gene-Related Peptide; Cricetinae; Electric Stimulation; Hibernation; Male; Mesocricetus; Neurokinin A; Substance P; Synaptic Transmission; Urinary Bladder; Vasoactive Intestinal Peptide

The pharmacological properties of non-adrenergic non-cholinergic (NANC) inhibitory neurotransmission were investigated in the rabbit choledocho-duodenal junction (CDJ), using the microelectrode and tension recording methods. L-NAME (10(-4) M) and apamin (5 X 10 (-6) M) suppressed NANC relaxation evoked by electrical field stimulation (EFS) in the presence of atropine and guanethidine (each 10(-6) M) to a similar extent (to about 40% of the initial control). However, combined application of L-NAME (10(-4) M) and apamin (5 X 10(-6) M) did not abolish it. EFS also evoked biphasic inhibitory junction potentials (IJPs) consisting of initial fast and slow sustained components in the presence of atropine and guanethidine (each 10(-6) M). Apamin (5 X 10(-8)-5 X 10(-6) M) dose-dependently suppressed the initial fast component by about 70%. In contrast, L-NAME (10(-4) M) did not affect either the amplitude of IJP or the resting membrane potential. PACAP-38 (> 10(-8) M) dose-dependently hyperpolarized the smooth muscle membrane of rabbit CDJ followed by a slow repolarization to the original level. After pretreatment with apamin (5 X 10(-7) M), PACAP-38 (10(-6) M) failed to evoke membrane hyperpolarization. During repolarization in the continued presence of PACAP-38, the amplitude of initial fast component of IJP was reduced to about 40-60% of control value, while that of the slow one was unaffected. A similar suppression of initial fast component of IJP (about 40% of the control value) also occurred after application of PACAP (6-38), a PACAP antagonist, or prolonged treatment with monoclonal antibodies to PACAP-27 or PACAP-38. Furthermore, the substantial part of residual fast IJP in the presence of PACAP (6-38) was suppressed by desensitization to alpha,beta-methylene ATP (10(-3) M). These results indicate that in rabbit CDJ NANC relaxation consists mainly of apamin- and L-NAME-sensitive components, which occur in a membrane potential dependent (through membrane hyperpolarization) and independent fashion, respectively. It has further been suggested that PACAP, together with a smaller contribution of ATP, may be involved as the principal apamin-sensitive transmitter in NANC relaxation of this muscle.

Topics: Adenosine Triphosphate; Animals; Autonomic Nervous System; Calcitonin Gene-Related Peptide; Common Bile Duct; Duodenum; Enzyme Inhibitors; In Vitro Techniques; Isometric Contraction; Male; Membrane Potentials; Neuropeptides; Neurotransmitter Agents; NG-Nitroarginine Methyl Ester; Nitric Oxide Synthase; Nitric Oxide Synthase Type I; Pituitary Adenylate Cyclase-Activating Polypeptide; Potassium Channels; Rabbits; Synaptic Transmission; Vasoactive Intestinal Peptide

1. Four putative neurotransmitters (serotonin, substance P, ATP (alpha-beta-methylene-ATP), and vasoactive intestinal peptide, VIP) of the non-adrenergic non-cholinergic (NANC) innervation were examined for their role in the NANC excitatory neurotransmission in channel catfish intestine after adrenergic and cholinergic blockade. 2. VIP at concentrations ranging from 10(-12)M to 10(-4)M did not produce either a relaxant or a contractile response in these segments. 3. Serotonin, substance P and alpha-beta-methyl-ATP produced contractile responses in a dose-dependent manner. Their EC50 values were 5 x 10(-7)M, 5 x 10(-9)M and 5 x 10(-9)M and 5 x 10(-6)M, respectively. 4. Electrical field stimulation of the intestinal segments produced a predominant excitatory response after complete blockade of adrenergic and cholinergic divisions, suggesting a predominant NANC excitatory innervation. 5. Three types of serotonin receptor antagonists, namely methiothepin (predominantly a 5-HT1 antagonist), ketanserin (a selective 5-HT2 antagonist), methysergide and cyproheptadine (predominantly 5-HT2 blockers) and metoclopramide (a selective 5-HT3 blocker) were tested for their effectiveness against serotonin and EFS-induced contractions. Methiothepin, methysergide, cyproheptadine and metoclopramide produced significant blockade of the response to serotonin, whereas only methiothepin and cyproheptadine produced blockade of EFS-induced response. 6. Three agents tested for substance P blockade, namely spantide, 4-11 fragment of substance P, and methysergide (also a serotonin blocker), did not produce significant inhibition of the response to either substance P or EFS. 7. Suramin at a dose that blocked the ED50 concentration of ATP did not produce a significant blockade of the response to EFS suggesting that ATP-involvement in the NANC-e neurotransmission is unlikely. 8. This study confirmed the involvement of serotonin in the expression of non-adrenergic non-cholinergic excitatory response of the channel catfish intestine.

Topics: Adenosine Triphosphate; Animals; Autonomic Nervous System; Electric Stimulation; Ictaluridae; In Vitro Techniques; Intestines; Serotonin; Substance P; Synaptic Transmission; Vasoactive Intestinal Peptide

In the guinea-pig proximal colon, 5-hydroxytryptamine (5-HT) relaxes the longitudinal muscle by stimulating neuronal 5-HT receptors, which induces the release of nitric oxide (NO). It was investigated whether the inhibitory neurotransmitters adenosine 5'-triphosphate (ATP) and/or vasoactive intestinal polypeptide (VIP) could be involved as well. Antagonists to block the contractile response to 5-HT via 5-HT2, 5-HT3 or 5-HT4 receptors were present throughout the experiments and methacholine was administered to precontract the strips. ATP, VIP and 5-HT induced concentration-dependent relaxations, in the case of 5-HT yielding a non-monophasic concentration-response curve. Tetrodotoxin (TTX; 300 nM), NG-nitro-L-arginine (L-NNA, 100 microM) and their combination did not inhibit the relaxations induced by VIP (up to 0.3 microM) or 0.3-3 microM ATP but reduced those by 10 microM ATP. Suramin (300 microM) strongly inhibited the relaxations to ATP and VIP. L-NNA and suramin also inhibited the relaxations to 5-HT. In the presence of L-NNA (100 microM), suramin did not significantly inhibit the relaxations to 5-HT. Suramin did not affect the relaxations to isoprenaline, nitroglycerin or exogenous NO (1 microM), demonstrating its specificity. Apamin (30 nM) inhibited both the relaxations to ATP (by 70-100%) and to 5-HT; relaxations to isoprenaline were partially inhibited, indicating a non-specific component in the inhibitory action of apamin. However, relaxations to exogenous VIP (up to 0.3 microM), NO (1 microM) and to nitroglycerin were not inhibited. In the presence of L-NNA (100 microM), apamin inhibited the relaxations to 5-HT only at 30 microM. alpha, beta-methylene-ATP (alpha, beta-Me-ATP; 100 microM) did not desensitize the responses to ATP. Reactive blue 2 affected the relaxations to isoprenaline at concentrations necessary to significantly inhibit the relaxations to ATP (i.e. from 10 microM onwards). Thus, it was not possible to test either alpha, beta-Me-ATP or reactive blue 2 against the relaxations to 5-HT. alpha-Chymotrypsin (0.015 mg.ml-1) and trypsin (0.005 mg.ml-1) almost abolished the relaxations to VIP, but did not affect those to isoprenaline and 5-HT. The VIP receptor antagonists [p-Cl-D-Phe6, Leu17]VIP (1 microM) and VIP10-28 (1 and 3 microM) did not affect the concentration-response curve to VIP and were hence not tested against 5-HT. Phosphoramidon (1 microM) had no effect on the relaxations to VIP or 5-HT.(ABSTRACT TRUNCATED AT 400 WORDS)

Topics: Adenosine Triphosphate; Animals; Apamin; Colon; Female; Guinea Pigs; In Vitro Techniques; Isoproterenol; Male; Muscle Relaxation; Muscle, Smooth; Nitric Oxide; Nitroglycerin; Serotonin; Suramin; Vasoactive Intestinal Peptide

A sucrose-gap technique was used to study the development of neuromuscular transmission in the taenia coli of fetal, 1- to 2-day-old, 3- to 4-week-old and 3-month-old guinea-pigs. In addition, the effects of exogenous, alpha,beta-methylene adenosine 5'-triphosphate (ATP), noradrenaline, vasoactive intestinal polypeptide (VIP) and carbachol were examined. Taking the response to a single pulse of electrical field stimulation as the index of a developed neuromuscular junction, it was apparent that the non-adrenergic inhibitory system arose before, and matured more quickly than, the cholinergic system. The inhibitory system was present by 8 weeks of gestation in some fetuses, but, in contrast, excitatory cholinergic transmission was not seen until birth. As evidenced by responses to carbachol, alpha,beta-methylene ATP and VIP, cholinergic, purinergic and VIP receptors were present on the smooth muscle at the earliest ages studied. No changes in sensitivity to these agents were noted throughout development, although in adults the level of the maximum responses increased.

Topics: Adenosine Triphosphate; Animals; Atropine; Autonomic Nervous System; Carbachol; Colon; Electric Stimulation; Electrophysiology; Female; Guinea Pigs; Male; Neuromuscular Junction; Norepinephrine; Parasympathetic Nervous System; Pregnancy; Synaptic Transmission; Vasoactive Intestinal Peptide

In the guinea-pig bladder, contractile responses to substance P (0.3 microM) and VIP (3 microM) were unaffected by P2-purinoceptor desensitization with alpha,beta-methylene ATP (3 X 10(-6) M), while the responses to stimulation of the non-cholinergic excitatory nerves (4-16 Hz) were abolished. The evidence presented suggests that ATP or a related purine nucleotide, and not VIP or substance P, is responsible for the non-cholinergic excitatory component of the nerve-mediated response.

Topics: Adenosine Triphosphate; Animals; Electric Stimulation; Female; Guinea Pigs; In Vitro Techniques; Isometric Contraction; Male; Neurons; Neurotransmitter Agents; Substance P; Urinary Bladder; Vasoactive Intestinal Peptide