BACKGROUND: The electrophysiological effects of purinergic receptor agonists, adenosine triphosphate(ATP) and adenosine were examined using conventional microelectrode technique in rat atrial muscle fibers under superfused with a normal or a simulated ischemic(hypoxic, hyperkalemic and acidotic) physiologic salt solution(PSS) in vitro. METHODS: Action potential parameters, such as maximal diastolic potential(MDP), action potential amplitude(APA), rate of phase 0 depolarization(dv/dtmax) and action potential duration(APD90) were measured in electrically paced, physiologic salt solution(Tyrode's) superfused left rat atrium. In the experiment of ischemic simulation in vitro, normal physiologic salt solutions(NPSS0 were modified(MPSS) and superfused in substitute for normal Tyrode's solution. To investigate the effects of purinergic receptor agonists, ATP or adenosine was added to the superfused tyrode's solutions(NPSS or MPSS) in molar concentration. RESULTS: Under superfused with normal PSS, ATP(10(-3), 10(-4)M) elicited slight hyperpolarization in MDP, and both ATP(10(-6)-10(-3)M) and adenosine(10(-6)-10(-3)M) shortened the duration of normal action potential in a dose-dependent manner. The other paramaters were not affected by the drugs. Superfusion with ischemic PSS caused reductions in MDP as well as APA, dv/dtmax and, especially, APD90. The effects produced by the initial 10 minutes of superfusion with ischemic PSS almost completely disappeared during a subsequent period of continued superfusion with normal PSS, but, those by the initial 20 min lasted in some degree. Both ATP(10(-4)M) and adenosine(10(-4)M) attenuated the reduction in the rate of phase 0 depolarization and the amplitude of the action potential amplitude produced by the ischemic PSS. CONCLUSION: Purinergic receptor agonists, ATP and adensoine, caused a concentration-dependent shortening of the action potential duration in rat atrial muscle fibers and they attenuated the reductions in the rate of phase 0 depolarization and action potential amplitude in fibers superfused with ischemic PSS.
Action Potentials ; Adenosine ; Adenosine Triphosphate ; Animals ; Ischemia ; Microelectrodes ; Molar ; Purinergic Agonists* ; Rats ; Receptors, Purinergic

BACKGROUND/AIMS: Adenosine is an endogenous modulator of nociception. Its role in visceral nociception, particularly in visceral hyperalgesia, has not been studied. The aim of this study was to determine the effects of adenosine receptor agonists in a model of visceral hyperalgesia. METHODS: The visceromotor response (VMR) in rats to colorectal distension (CRD; 80 mmHg, 20 seconds) was quantified by electromyographic recordings from the abdominal musculature. Three hours after the intracolonic administration of zymosan (25 mg/mL, 1 mL), VMRs to CRD were measured before and after either subcutaneous or intrathecal administration of an adenosine receptor agonist. RESULTS: Subcutaneous injection of 5'-N-ethylcarboxyamidoadenosine (NECA; an A1 and A2 receptor agonist), R(-)-N6-(2-phenylisopropyl)-adenosine (R-PIA; a selective A1 receptor agonist), or CGS-21680 hydrochloride (a selective A2a receptor agonist) dose-dependently (10-100 mg/kg) attenuated the VMR to CRD, although hindlimb weakness occurred at the higher doses tested. Intrathecal administration of NECA or R-PIA dose-dependently (0.1-1.0 microgram/kg) decreased the VMR, whereas CGS-21680 hydrochloride was ineffective over the same concentration range. Higher intrathecal doses of the A1/A2 receptor agonist NECA produced motor weakness. CONCLUSIONS: Adenosine receptor agonists are antihyperalgesic, but also produce motor weakness at high doses. However, activation of the spinal A1 receptor significantly attenuates the VMR to CRD without producing motor weakness.
Adenosine ; Adenosine-5'-(N-ethylcarboxamide) ; Animals ; Hindlimb ; Hyperalgesia ; Injections, Subcutaneous ; Nociception ; Purinergic P1 Receptor Agonists ; Rats ; Receptors, Purinergic P1 ; Zymosan

PURPOSE: To evaluate changes in the tear meniscus area and tear meniscus height over time in patients with dry eye syndrome, using anterior segment spectral-domain optical coherence tomography after the instillation of 3% diquafosol ophthalmic solution. METHODS: Sixty eyes from 30 patients with mild to moderate dry eye syndrome were included. Tear meniscus images acquired by anterior segment spectral-domain optical coherence tomography were analyzed using National Institutes of Health's image-analysis software (ImageJ 1.44p). Tear meniscus area and tear meniscus height were measured at baseline, 5 minutes, 10 minutes, and 30 minutes after instillation of a drop of diquafosol in one eye and normal saline in the other eye. Changes in ocular surface disease index score, tear film break-up time, corneal staining score by Oxford schema, and meibomian expressibility were also evaluated at baseline, and after 1 week and 1 month of a diquafosol daily regimen. RESULTS: Sixty eyes from 30 subjects (mean age, 29.3 years; 8 men and 22 women) were included. In eyes receiving diquafosol, tear volume was increased at 5 and 10 minutes compared with baseline. It was also higher than saline instilled eyes at 5, 10, and 30 minutes. Changes in tear volume with respect to baseline were not statistically different after the use of diquafosol for 1 month. Ocular surface disease index score, tear film break-up time, and Oxford cornea stain score were significantly improved after 1 week and 1 month of daily diquafosol instillation, but meibomian expressibility did not change. CONCLUSIONS: Topical diquafosol ophthalmic solution effectively increased tear volume for up to 30 minutes, compared to normal saline in patients with dry eye syndrome.
Academies and Institutes ; Cornea ; Dry Eye Syndromes* ; Humans ; Male ; Ophthalmic Solutions ; Purinergic P2Y Receptor Agonists ; Tears* ; Tomography, Optical Coherence*

BACKGROUND AND OBJECTIVES: Puringeric receptors and their agonists like uridine-5-triphosphate (UTP) and adenosine triphosphate (ATP), regulate mucin secretion in middle ear epithelial cells. In the present study, we examined the effects of purinergic agonists on Ca2+ influx ([Ca2+]i ) in normal human middle ear epithelial (NHMEE) cells. We also examined the effect of caffeine, an inositol 1, 4, 5-triphosphate (IP3) inhibitor, on UTP induced [Ca2+]i and mucin secretion in NHMEE cells. MATERIALS AND METHOD: NHMEE cells were stimulated with various purinergic agonists, such as UTP, and [Ca2+]i was measured using a miniature double perfusion chamber. UTP-induced mucin secretion was quantitated by immunoblotting assay. RESULTS: The determined order of purinergic agonist potency with respect to [Ca2+]i was ATP=UTP>2-MeSATP>ADP>> adenosine. UTP-induced mucin secretion was inhibited when the intracellular Ca2+ was removed with 2-bis (2-aminophenoxy)ethane-N, N, N', N'-tetraacetic acid-acetoxymethyl ester. Caffeine suppressed UTP-induced [Ca2+]i, and but inhibited UTPinduced and constitutional mucin secretion. CONCLUSION: Our results suggest that caffeine may have a therapeutic effect in mucoid otitis media by suppressing mucin secretion.
Adenosine ; Adenosine Triphosphate ; Caffeine* ; Calcium ; Ear, Middle* ; Epithelial Cells* ; Humans* ; Immunoblotting ; Inositol ; Mucins* ; Otitis Media ; Perfusion ; Purinergic Agonists ; Uridine Triphosphate

BACKGROUND: A Nerve ligation injury may produce a pain syndrome that includes tactile allodynia. Reversal effects on tactile allodynia have been demonstrated after an intrathecal administration of adenosine analogues or morphine. Adenosine receptor agonists have been known to have antinociceptive and antiallodynic effects in many animal and human studies. We examined the drug interactions between morphine and adenosine agonists in a rat model of a nerve ligation injury. METHODS: Male Sprague Dawley rats were prepared with a tight ligation of the left lumbar 5 th and 6 th spinal nerves and chronic lumbar intrathecal catheter implantation for drug administration. We measured the tactile allodynia by applying von Frey filaments ipsilateral to the lesioned hindpaw. Thresholds for paw withdrawal were assessed. Morphine (1 - 30ng), adenosine (1 - 30ng) and R-PIA (0.1 - 10ng) were administered to obtain the dose-response curves and the 50% effective dose (ED50). Fractions of ED50 values were administered to establish the ED50 of drug combinations. Drug interactions were evaluated by the fractional and isobolographic analyses. Allodynic thresholds for left lesioned hindpaw withdrawal to the von Frey hairs test were assessed and converted to % maximal possible effect (%MPE). RESULTS: The antiallodynic effect of morphine, adenosine, and R-PIA were produced in a dose dependent manner. The antiallodynic effects of combinations showed a similar pattern. Isobolographic analysis revealed a synergistic interaction for the morphine-R-PIA combination but not for the morphine-adenosine combination. However, fractional analysis produced a synergistic result for two combination groups. CONCLUSIONS: The results demonstrated that intrathecal co-administration of adenosine A1 receptors agonist and morphine showed the synergistic effect on nerve ligation injury induced allodynia.
Adenosine* ; Animals ; Catheters ; Drug Combinations ; Drug Interactions ; Hair ; Humans ; Hyperalgesia ; Injections, Spinal ; Ligation* ; Male ; Models, Animal ; Morphine* ; Purinergic P1 Receptor Agonists* ; Rats* ; Rats, Sprague-Dawley ; Receptor, Adenosine A1 ; Receptors, Purinergic P1* ; Spinal Nerves

An important property of the intestine is the ability to secrete fluid. The intestinal secretion is regulated by a number of substances including vasoactive intestinal peptide (VIP), ATP and different inflammatory mediators. One of the most important secretagogues is adenosine during inflammation. However, the controversy concerning the underlying mechanism of adenosine-stimulated Cl- secretion in intestinal epithelial cells still continues. To investigate the effect of adenosine on Cl- secretion and its underlying mechanism in the rabbit colon mucosa, we measured short circuit current (ISC) under automatic voltage clamp with DVC-1000 in a modified Ussing chamber. Adenosine, when added to the basolateral side of the mucosa, increased ISC in a dose-dependent manner. The adenosine-stimulated ISC response was abolished when Cl- in the bath solution was replaced completely with gluconate. In addition, the ISC response was inhibited by a basolateral Na-K-Cl cotransporter blocker, bumetanide, and by apical Clchannel blockers, dephenylamine-2-carboxylate (DPC), 5-nitro-2-(3-phenyl-propylamino)-benzoate (NPPB), glibenclamide. Amiloride, an epithelial Na+ channel blocker, and 4,4-diisothiocyanato-stilbene-2,2-disulphonate (DIDS), a Ca2+-activated Cl- channel blocker, had no effect. In the mucosa pre-stimulated with forskolin, adenosine did not show any additive effect, whereas carbachol resulted in a synergistic potentiation of the ISC response. The adenosine response was inhibited by 10 micrometer H-89, an inhibitor of protein kinase A. These results suggest that the adenosine-stimulated ISC response is mediated by basolateral to apical Cl- secretion through a cAMP-dependent Cl- channel. The rank order of potencies of adenosine receptor agonists was 5'-(N-ethylcarboxamino)adenosine(NECA) > N6-(R-phenylisopropyl)adenosine(R-PIA)>2-(p-(2-carbonylethyl)-phenyl-et hylamino)-5'-N-ethylcarboxaminoadenosine(CGS21680). From the above results, it can be concluded that adenosine interacts with the A2b adenosine receptor in the rabbit colon mucosa and a cAMP-dependent signalling mechanism underlies the stimulation of Cl- secretion.
Adenosine ; Adenosine Triphosphate ; Amiloride ; Baths ; Bumetanide ; Carbachol ; Colforsin ; Colon* ; Cyclic AMP-Dependent Protein Kinases ; Epithelial Cells ; Glyburide ; Inflammation ; Intestinal Secretions ; Intestines ; Mucous Membrane* ; Purinergic P1 Receptor Agonists ; Receptors, Purinergic P1 ; Vasoactive Intestinal Peptide

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.
Adenosine Triphosphate ; Animals ; Atropine ; Bethanechol ; Choline ; Cholinergic Agonists ; Cholinesterases ; Ganglion Cysts ; Hemicholinium 3 ; Hexamethonium ; Myography ; Nerve Endings ; Neural Conduction ; Physostigmine ; Rats* ; Receptors, Purinergic ; Tetrodotoxin ; Urinary Bladder*

BACKGROUND: Nerve ligation injury in rats produces a pain syndrome that includes mechanical allodynia. Intrathecal administration of cholinesterase inhibitors or adenosine receptor agonists have anti-allodynic effects in this model. Therefore, we tested the interaction between intrathecal neostigmine and N(6)-cyclohexyladenosine (CHA) in a rat behavioral model of neuropathic pain. METHODS: Male Sprague-Dawley rats were prepared with tight ligation of the spinal nerves for producing allodynia and with a lumbar intrathecal catheter for drug administration. Allodynia thresholds for hindpaw withdrawal against mechanical stimuli were assessed and converted to percent maximal possible effect. Neostigmine (0.3-10 microgram) and CHA (0.03-3 microgram) were administered to obtain the dose-response curves and the 50% effective dose (ED(50)). Equal fractions (1/2, 1/4 and 1/8 ED(50)s) of the two drugs were administered to establish the ED(50) of neostigmine-CHA combination. Side effects were also assessed. The drug interaction was evaluated by isobolographic and fractional analyses. RESULTS: Neostigmine, CHA, and the neostigmine-CHA combination dose-dependently produced anti-allodynia effects. Side effects such as sedation and motor weakness were similar in the three groups. In the isobolographic analysis, the experimental ED(50) for the combination of neostigmine-CHA lay far below and to the left of the theoretical additive line. Fractional analysis indicated that the total combination fraction of the two drugs was 0.39. CONCLUSIONS: Intrathecal co-administration of neostigmine and CHA showed a synergistic anti-allodynia effect.
Adenosine ; Animals ; Catheters ; Cholinesterase Inhibitors ; Drug Interactions ; Humans ; Hyperalgesia ; Injections, Spinal ; Ligation ; Male ; Neostigmine ; Neuralgia ; Purinergic P1 Receptor Agonists ; Rats ; Rats, Sprague-Dawley ; Spinal Nerves

OBJECTIVETo study the therapeutic efficacy and mechanism of adenosine receptor agonist N(6)-cyclopentyladenosine (CPA) against acute dichlorvos poisoning.

METHODSSoon after a certain doses of dichlorvos were given to mice and rats by gastrogavage, physiological saline, CPA, atropine or pralidoxime chloride were administered to different groups. Toxic signs and survival rate were recorded and cholinesterase (ChE) activities and acetylcholine (ACh) concentrations in whole blood were determined in treatment group with CPA and non-treatment group after dichlorvos was given to rats by gastrogavage.

RESULTS(1) The alleviated and delayed appearance of toxic signs as well as obvious prolongation of survival time was observed in CPA treatment group compared with non-treatment group. (2) ChE activities in both treatment group with CPA [(0.49 +/- 0.05) U/ml] and non-treatment group [(0.52 +/- 0.04) U/ml] were significantly lower than that [(1.56 +/- 0.15) U/ml] of the control group (P < 0.01), but there was no significant difference between treatment group and non-treatment group (P > 0.05). (3) ACh concentration [(204.24 +/- 20.48) microg/ml] in whole blood of treatment group with CPA was significantly lower than that [(230.91 +/- 25.61) microg/ml] of non-treatment group (P < 0.05).

CONCLUSIONCPA has therapeutic efficacy against acute dichlorvos poisoning, which is probably through the decrease in ACh concentration.

Acetylcholine ; blood ; Acute Disease ; Adenosine ; analogs & derivatives ; therapeutic use ; Animals ; Cholinesterases ; blood ; Dichlorvos ; poisoning ; Insecticides ; poisoning ; Male ; Mice ; Mice, Inbred Strains ; Poisoning ; blood ; drug therapy ; Purinergic P1 Receptor Agonists ; Rats ; Rats, Sprague-Dawley

In order to investigate the role of non-adrenergic non-cholinergic nerves in regulating mechanical and electrical activity of gastric circular smooth muscle, the effects of ATP and its analogues on gastric motility and electrical activities were observed in guinea-pig. In organ bath system, isometric force of the circular smooth muscle of guinea-pig gastric antrum was measured. Electrical activity of the muscle was recorded by using intracellular microelectrode. Electrical and mechanical activities were recorded by chart recorder. ATP and 2-MeSATP potentiated the mechanical activity but did not affect electrical activity in gastric circular smooth muscle. ATP and 2-MeSATP-induced contraction was effectively blocked by nonselective P2y-purinoceptor antagonist, reactive-blue-2 and suramin, but ATP-induced contraction was not blocked by alpha,beta-MeATP-induced desensitization of P2x-purinoceptors. However, alpha,beta-MeATP, P2x-purinoceptor agonist, attenuated slow waves with membrane hyperpolarization and inhibited contraction. The relaxation by beta,gamma-MeATP was blocked by alpha,beta-MeATP-induced desensitization of P2x-purinoceptors. ATP-induced contraction was blocked by external calcium-free, but not by nicardipine, a L-type calcium channel blocker. Indomethacin, a nonselective cyclooxygenase inhibitor, did not block ATP-induced contraction. The results suggest that: (1) ATP- and analogues-induced contraction is mediated by P2y-purinoceptor, whereas alpha,beta-MeATP-induced relaxation by P2x-purinoceptor in guinea-pig gastric antral circular smooth muscle. (2) ATP-induced contraction is dependent on extracellular calcium, but Ca2+ entry is not mediated by L-type calcium channel. (3) Prostaglandins are not involved in ATP- and analogue-induced contraction of gastric circular smooth muscle in guinea-pigs, and alpha,beta-MeATP-induced relaxation is related to membrane hyperpolarization.
Adenosine Triphosphate ; analogs & derivatives ; pharmacology ; Animals ; Electrophysiology ; Guinea Pigs ; In Vitro Techniques ; Microelectrodes ; Muscle Contraction ; drug effects ; physiology ; Muscle, Smooth ; drug effects ; physiology ; Purinergic Agonists ; Pyloric Antrum ; drug effects ; physiology ; Thionucleotides ; pharmacology