On the mechanism of histaminergic inhibition of glutamate release in the rat dentate gyrus.
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1. Histaminergic depression of excitatory synaptic transmission in the rat dentate gyrus was investigated using extracellular and whole-cell patch-clamp recording techniques in vitro. 2. Application of histamine (10 microM, 5 min) depressed synaptic transmission in the dentate gyrus for 1 h. This depression was blocked by the selective antagonist of histamine H3 receptors, thioperamide (10 microM). 3. The magnitude of the depression caused by histamine was inversely related to the extracellular Ca2+ concentration. Application of the N-type calcium channel blocker omega-conotoxin (0. 5 or 1 microM) or the P/Q-type calcium channel blocker omega-agatoxin (800 nM) did not prevent depression of synaptic transmission by histamine. 4. The potassium channel blocker 4-aminopyridine (4-AP, 100 microM) enhanced synaptic transmission and reduced the depressant effect of histamine (10 microM). 4-AP reduced the effect of histamine more in 2 mM extracellular calcium than in 4 mM extracellular calcium. 5. Histamine (10 microM) did not affect the amplitude of miniature excitatory postsynaptic currents (mEPSCs) and had only a small effect on their frequency. 6. Histaminergic depression was not blocked by an inhibitor of serine/threonine protein kinases, H7 (100 microM), or by an inhibitor of tyrosine kinases, Lavendustin A (10 microM). 7. Application of adenosine (20 microM) or the adenosine A1 agonist N6-cyclopentyladenosine (CPA, 0.3 microM) completely occluded the effect of histamine (10 microM). 8. We conclude that histamine, acting on histamine H3 receptors, inhibits glutamate release by inhibiting presynaptic calcium entry, via a direct G-protein-mediated inhibition of multiple calcium channels. Histamine H3 receptors and adenosine A1 receptors act upon a common final effector to cause presynaptic inhibition. (+info)
Inhibition by adenosine receptor agonists of synaptic transmission in rat periaqueductal grey neurons.
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1. The actions of selective adenosine A1 and A2 receptor agonists were examined on synaptic currents in periaqueductal grey (PAG) neurons using patch-clamp recordings in brain slices. 2. The A1 receptor agonist 2-chloro-N-cyclopentyladenosine (CCPA), but not the A2 agonist, 2-p-(2-carboxyethyl)phenethylamino-5'-N-ethylcarboxamidoadenosine (CGS21680), inhibited both electrically evoked inhibitory (eIPSCs) and excitatory (eEPSCs) postsynaptic currents. The actions of CCPA were reversed by the A1 receptor antagonist 8-cyclopentyl-1, 3-dipropylxanthine (DPCPX). 3. In the absence or presence of forskolin, DPCPX had no effect on eIPSCs, suggesting that concentrations of tonically released adenosine are not sufficient to inhibit synaptic transmission in the PAG. 4. CCPA decreased the frequency of spontaneous miniature action potential-independent IPSCs (mIPSCs) but had no effect on their amplitude distributions. Inhibition persisted in nominally Ca2+-free, high Mg2+ solutions and in 4-aminopyridine. 5. The CCPA-induced decrease in mIPSC frequency was partially blocked by the non-selective protein kinase inhibitor staurosporine, the specific protein kinase A inhibitor 8-para-chlorophenylthioadenosine-3',5'-cyclic monophosphorothioate (Rp-8-CPT-cAMPS), and by 8-bromoadenosine cyclic 3',5' monophosphate (8-Br-cAMP). 6. These results suggest that A1 adenosine receptor agonists inhibit both GABAergic and glutamatergic synaptic transmission in the PAG. Inhibition of GABAergic transmission is mediated by presynaptic mechanisms that partly involve protein kinase A. (+info)
Brain blood flow and blood pressure during hypoxia in the epaulette shark Hemiscyllium ocellatum, a hypoxia-tolerant elasmobranch.
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The key to surviving hypoxia is to protect the brain from energy depletion. The epaulette shark (Hemiscyllium ocellatum) is an elasmobranch able to resist energy depletion and to survive hypoxia. Using epi-illumination microscopy in vivo to observe cerebral blood flow velocity on the brain surface, we show that cerebral blood flow in the epaulette shark is unaffected by 2 h of severe hypoxia (0.35 mg O2 l-1 in the respiratory water, 24 C). Thus, the epaulette shark differs from other hypoxia- and anoxia-tolerant species studied: there is no adenosine-mediated increase in cerebral blood flow such as that occurring in freshwater turtles and cyprinid fish. However, blood pressure showed a 50 % decrease in the epaulette shark during hypoxia, indicating that a compensatory cerebral vasodilatation occurs to maintain cerebral blood flow. We observed an increase in cerebral blood flow velocity when superfusing the normoxic brain with adenosine (making sharks the oldest vertebrate group in which this mechanism has been found). The adenosine-induced increase in cerebral blood flow velocity was reduced by the adenosine receptor antagonist aminophylline. Aminophylline had no effect upon the maintenance of cerebral blood flow during hypoxia, however, indicating that adenosine is not involved in maintaining cerebral blood flow in the epaulette shark during hypoxic hypotension. (+info)
Role of renal medullary adenosine in the control of blood flow and sodium excretion.
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This study determined the levels of adenosine in the renal medullary interstitium using microdialysis and fluorescence HPLC techniques and examined the role of endogenous adenosine in the control of medullary blood flow and sodium excretion by infusing the specific adenosine receptor antagonists or agonists into the renal medulla of anesthetized Sprague-Dawley rats. Renal cortical and medullary blood flows were measured using laser-Doppler flowmetry. Analysis of microdialyzed samples showed that the adenosine concentration in the renal medullary interstitial dialysate averaged 212 +/- 5.2 nM, which was significantly higher than 55.6 +/- 5.3 nM in the renal cortex (n = 9). Renal medullary interstitial infusion of a selective A1 antagonist, 8-cyclopentyl-1,3-dipropylxanthine (DPCPX; 300 pmol. kg-1. min-1, n = 8), did not alter renal blood flows, but increased urine flow by 37% and sodium excretion by 42%. In contrast, renal medullary infusion of the selective A2 receptor blocker 3, 7-dimethyl-1-propargylxanthine (DMPX; 150 pmol. kg-1. min-1, n = 9) decreased outer medullary blood flow (OMBF) by 28%, inner medullary blood flows (IMBF) by 21%, and sodium excretion by 35%. Renal medullary interstitial infusion of adenosine produced a dose-dependent increase in OMBF, IMBF, urine flow, and sodium excretion at doses from 3 to 300 pmol. kg-1. min-1 (n = 7). These effects of adenosine were markedly attenuated by the pretreatment of DMPX, but unaltered by DPCPX. Infusion of a selective A3 receptor agonist, N6-benzyl-5'-(N-ethylcarbonxamido)adenosine (300 pmol. kg-1. min-1, n = 6) into the renal medulla had no effect on medullary blood flows or renal function. Glomerular filtration rate and arterial pressure were not changed by medullary infusion of any drugs. Our results indicate that endogenous medullary adenosine at physiological concentrations serves to dilate medullary vessels via A2 receptors, resulting in a natriuretic response that overrides the tubular A1 receptor-mediated antinatriuretic effects. (+info)
Endogenous interstitial adenosine in isolated myenteric neural networks varies inversely with prevailing PO2.
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Isolated myenteric ganglion networks were used in a perifusion protocol to characterize the response of interstitial adenosine levels to changes in prevailing PO2. The biological activity of such adenosine was assessed using inhibition of release of substance P (SP) as a functional measure of adenosine activity, and the effect of altered O2 tension on both spontaneous and elevated extracellular K+ concentration-evoked SP release from networks was determined over a range of PO2 values from hypoxic (PO2 = 54 mmHg) to hyperoxic (PO2 = 566 mmHg). Release of SP was found to be sensitive to PO2, and a linear graded relationship was obtained. Perifusion in the additional presence of the adenosine A1-receptor-selective antagonist 1,3-dipropyl-8-cyclopentylxanthine (DPCPX) revealed considerable adenosinergic inhibition with an inverse exponential relationship and hyperoxic threshold PO2. Disinhibition of evoked SP release by DPCPX in the absence of TTX was double that observed in its presence, indicating a neural source for some of the adenosine released during hypoxia. A postulated neuroprotective role for adenosine is consistent with the demonstrated relationship between interstitial adenosine and prevailing O2 tension. (+info)
Natriuretic and diuretic actions of a highly selective adenosine A1 receptor antagonist.
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The natriuretic and diuretic action of a highly selective adenosine A1 receptor (A1AdoR) antagonist, 1,3-dipropyl-8-[2-(5,6-epoxy)norbornyl]xanthine (CVT-124), was investigated in anesthetized rats. CVT-124 (0.1 to 1 mg/kg) caused dose-dependent increases in urine flow and fractional and absolute sodium excretion of by six- to 10-fold and, at 0.1 mg/kg, increased the GFR (1.6+/-0.1 to 2.5+/-0.2 ml/min; P<0.01). There were no changes in BP or heart rate. CVT-124 reduced absolute proximal reabsorption (26+/-3 to 20+/-2 nl/min; P<0.05) despite unchanged proximally measured, single-nephron GFR (SNGFR) (42+/-5 to 44+/-4 nl/min; NS) and thereby decreased fractional proximal reabsorption (60+/-3 to 46+/-4%; P<0.05). Despite increasing distal tubular fluid flow rate (5.4+/-0.7 to 9.7+/-0.9 nl/min; P<0.001), it reduced the proximal-distal difference in SNGFR (before: 9.4+/-1.0 versus during CVT-124: 4.6+/-1.5 nl/min; P<0.01), suggesting that it had blunted the effects of the macula densa on SNGFR. Direct measurements of maximal tubuloglomerular feedback (TGF) responses were made from proximal stop flow pressure (PSF) during orthograde loop perfusion from the proximal tubule with artificial tubular fluid at 40 nl/min. TGF was blunted by intravenous CVT-124 (0.5 mg/kg; deltaPSF with vehicle: 8.3+/-0.6 versus CVT-124: 6.5+/-0.3 mm Hg; n = 9; P<0.01). In conclusion, A1AdoR blockade reduces proximal reabsorption and uncouples it from glomerular filtration. It increases distal delivery of fluid yet does not activate a macula densa-dependent fall in SNGFR because it blunts the TGF response. Natriuresis accompanied by blockade of proximal glomerulotubular balance and TGF characterizes a new class of diuretic drugs. (+info)
Retinal ischemic preconditioning in the rat: requirement for adenosine and repetitive induction.
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PURPOSE: A brief period of ischemia can induce a remarkably complete state of ischemic tolerance in the retina, a phenomenon known as ischemic preconditioning (IPC). The mechanisms of IPC were studied in the rat retina by examining the role of adenosine as a possible mediator and determining whether IPC protection could be induced more than once in the same rat. METHODS: Retinal ischemia was produced for 60 minutes in ketamine-xylazine-anesthetized Sprague-Dawley rats, and recovery was measured using electroretinography. Twenty-four hours earlier, the IPC stimulus of 5 minutes of ischemia was applied. To test the role of adenosine as a mediator of IPC, the selective adenosine A1 receptor antagonist 8-cyclopentyl-1,3-dipropylxanthine (DPCPX; 0.45 mg/kg, 2.25 mg/kg, or 4.5 mg/kg), the A2a antagonist 8-(3-chlorostyryl)caffeine (CSC; 0.1 mg/kg or 1.0 mg/kg), or their cyclodextrin vehicle were administered 15 minutes before IPC. To examine whether exogenous adenosine administration could mimic IPC, animals received intravitreal injections of the adenosine A1 receptor stimulant adenosine amine congener (ADAC) or the A2a stimulant CGS21680, followed by ischemia 24 hours later. To test the hypothesis that IPC could be induced repeatedly without loss of protection, rats were divided to receive IPC or sham IPC, followed 10 days later by IPC or a sham procedure, and 24 hours later by 60 minutes of ischemia. RESULTS: Adenosine A1 receptor blockade with 4.5 mg/kg DPCPX administered intraperitoneally (IP) before or immediately after 5 minutes of ischemia completely blocked IPC protection, whereas lower doses resulted in partial blockade. CSC at the lowest dose (0.1 mg/kg) had no significant effect on IPC's protective effect, whereas partial blockade was found with 1.0 mg/kg CSC. A1 or A2a receptor stimulation produced partial but significant mimicking of IPC protection, effects that were antagonized by DPCPX or CSC. Ischemic preconditioning applied twice, separated by 10 days, and followed by 60 minutes of ischemia 24 hours after the second IPC stimulus, resulted in nearly identical recovery of function after ischemia compared with IPC performed one time. CONCLUSIONS: Adenosine, acting through the A1 and A2a receptors, is a critical component in the induction of ischemic tolerance after preconditioning in the retina. The neuroprotective effects of IPC in the retina are lost over time but may be reinduced by subsequent application of the IPC stimulus. (+info)
P2 purinoceptors contribute to ATP-induced inhibition of L-type Ca2+ current in rabbit atrial myocytes.
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OBJECTIVE: Adenine compounds, including adenosine-5'-triphosphate (ATP) and adenosine (Ado), exert inhibitory effects on myocardium via P1 (subtype A1) purinoceptors. However, ATP per se is a potent activator of P2 purinoceptors. Our aim was to elucidate the respective roles of P1 and P2 purinoceptors in the actions of ATP on L-type calcium current (ICa) in rabbit atrial cells. METHODS AND RESULTS: A whole cell clamp technique was used to record ICa in single atrial cells from the rabbit heart. ATP (0.1 mumol/1-3 mmol/l) produced an inhibitory effect on ICa prestimulated by isoproterenol (ISO, 30 nmol/l), even in the presence of Ado (1 mmol/l). Both 1,3-dipropyl-8-cyclopentylxanthine (A1 blocker) and suramin (P2 blocker) partially blocked the ATP-induced inhibition of ICa, while their co-application nearly completely abolished the effect of ATP. ATP-gamma S (30 mumol/l) inhibited ISO-stimulated ICa significantly, and this inhibition was completely blocked by suramin. alpha, beta-Methylene-ADP, an inhibitor of hydrolysis of AMP to Ado, eliminated the suramin-resistant component of ICa inhibition by ATP. Pretreatment with pertussis toxin (PTX) abolished the ATP inhibition of ICa. Both intracellular dialysis with 8Br cAMP and the application of forskolin plus 3-isobutyl-1-methylxanthine also eliminated the effect of ATP. CONCLUSIONS: Both P1 and P2 purinoceptors are involved in the ATP inhibition of ISO-stimulated ICa in rabbit atrial cells. The P1 stimulation by ATP results from hydrolysis of ATP to Ado. Both the P2- and the P1-mediated effects of ATP and Ado, respectively. involve a PTX-sensitive and cAMP-dependent pathway. (+info)