Evidence for an endogenous cAMP-adenosine pathway in the rat kidney

In the rat kidney, exogenous adenosine-3'-5'-monophosphate (cAMP) is converted to adenosine via the metabolism of cAMP to adenosine-5'-monophosphate by phosphodiesterase and adenosine-5'-monophosphate to adenosine by 5'-nucleotidase. Our purpose was to investigate whether in the rat kidney adenosine is synthesized from endogenous cAMP via the same pathway. Rat kidneys were perfused with Tyrode's solution, and stabilized for 3 hr to minimize basal renal purine secretion. In control experiments (n = 6), the renal venous secretion rate of adenosine, inosine, hypoxanthine and Sigmapurines (adenosine + inosine + hypoxanthine) did not change over the two 10-min experimental periods. In contrast, the beta adrenoceptor agonist (+/-)-isoproterenol (1 and 10 microM added to the perfusate) caused a significant (1-factor analysis of variance with repeated measures; n = 31) increase in the renal venous secretion of adenosine (P <.0001), inosine (P <.0007), hypoxanthine (P <.0007) and Sigmapurines (P <.0001) as measured by high-performance liquid chromatography with ultraviolet detection. The Sigmapurines was the most discriminating index of isoproterenol-induced changes in purine release, and the renal venous secretion of Sigmapurines was significantly (2-factor analysis of variance with repeated measures) attenuated by inhibition of beta adrenoceptors with propranolol (.1 microM, n = 6; P <.05), phosphodiesterase with 3-isobutyl-1-methylxanthine (1 mM, n = 5; P <.002) and 5'-nucleotidase with alpha, beta-methyleneadenosine-5'-diphosphate (0.1 mM, n = 5; P <.03). Our data indicate that activation of beta adrenoceptors increases purine biosynthesis in the rat kidney via a mechanism that involves phosphodiesterase and 5'-nucleotidase. These results support the existence of an endogenous cAMP-adenosine pathway in the rat kidney.     

Effects of ischemia, preconditioning, and adenosine deaminase inhibition on interstitial adenosine levels and infarct size

In order to examine the relationship between local adenosine concentrations before, during, and after ischemia and the extent of ischemic myocardial damage, measurements of interstitial fluid (ISF) nucleosides were made using microdialysis probes implanted in the ischemic region of isoflurane anesthetized Micropigs undergoing 60' coronary artery occlusion (CAO) and 3 h of reperfusion (REP). Nucleoside concentrations in the dialysate collected from the microdialysis probes were used as an index of ISF levels. Dialysate nucleoside concentrations (ADO, inosine and hypoxanthine), myocardial infarct size, and myocardial blood flow (MBF) were determined in control animals (n = 6), animals preconditioned with a single 10' cycle of CAO and REP (PC, n = 6), and those treated with the adenosine deaminase inhibitor pentostatin (n = 6, 0.2 mg/Kg i.v. 30' prior to CAO). The brief PC occlusion resulted in a transient but significant increase in dialysate ADO (6.7 +/- 1.8 microM vs. 0.67 +/- 0.1 microM at baseline). Pentostatin administration had no significant effect on either dialysate nucleosides or MBF at baseline. During the 60' CAO, dialysate ADO increased in control animals. In PC animals, however, dialysate ADO during CAO was lower than control. Pretreatment with pentostatin resulted in a six-fold augmentation in dialysate ADO during the 60 min CAO when compared to the control values (110.62 +/- 30.2 microM vs. 16.31 +/- 2.1 microM at 60 min of ischemia). Pentostatin also resulted in a significant reduction in the accumulation of inosine and hypoxanthine, indicating inhibition of adenosine deaminase activity. There were no significant differences in MBF between groups at any time point. Following 3 h REP, infarct size was 35.4 +/- 5.5%, 8.1 +/- 1.5% and 8.3 +/- 1.8% of the region at risk in control, PC, and pentostatin groups, respectively. These data suggest that marked increase in ISF ADO during CAO, may be as effective in reducing INF as a modest increase in ISF ADO prior to prolonged CAO.     

Comparison of interstitial fluid and coronary venous adenosine levels in in vivo porcine myocardium

There are numerous reports of interstitial fluid (ISF) and coronary venous adenosine measurements in isolated perfused hearts. This study was designed to simultaneously compare ISF and coronary venous adenosine concentrations during various interventions in in vivo porcine myocardium. In anesthetized, open-chest pigs, ISF adenosine, inosine, and hypoxanthine were sampled with cardiac microdialysis. Coronary sinus or venous purines were sampled with a metabolism-stop solution. During basal conditions, ISF adenosine was greater than coronary venous adenosine, but vascular inosine and hypoxanthine were greater than corresponding ISF levels. Dobutamine (20 micrograms/kg/min, i.v.) and systemic hypoxia produced three- and two-fold increases in ISF adenosine, but had no significant effect on coronary sinus adenosine concentration. Hypoxia, but not dobutamine, increased coronary sinus total purines 50%. In contrast to these interventions, intracoronary adenosine infusion (0.5-50 micrograms/kg/min) was associated with significantly greater coronary venous adenosine concentrations than ISF levels. Only during a coronary artery occlusion/reperfusion protocol were ISF and coronary venous adenosine concentrations comparable. The results of this study thus provide in vivo evidence of the powerful endothelial and red blood cell metabolic barriers to both exogenous and endogenous adenosine. These results also illustrate the differences in adenosine concentrations in the ISF and vascular spaces.     

In vivo and in vitro developmental toxicity in LPS-induced zinc-deficient rabbits

Adenosine, produced from the decomposition of adenosine triphosphate, is believed to provide protective effects during ischemia. On the other hand, adenosine metabolites may serve as precursors for oxygen free radical formation. The time course of formation of adenosine and its purine metabolites was studied during retinal ischemia in rats. Concentrations of adenosine and its purine nucleoside metabolites inosine, hypoxanthine, and xanthine in the retina-choroid of ketamine/xylazine-anesthetized rats were measured during retinal ischemia using high performance liquid chromatography. Quantitative measurements were made possible in the small tissue mass through the use of internal standards. Ischemia was induced by ligation of the central retinal artery. In each rat, one eye was ischemic while the other served as a non-ischemic control. Eyes were frozen in situ at 1, 5, 10, 20, 30, 60, and 120 min of ischemia. The retina-choroid was then removed from the frozen eyes and analysed. Significant increases in the concentrations of adenosine, inosine, and hypoxanthine in ischemic compared to control retina-choroid were detectable within 1 to 5 min of the onset of ischemia, and within 10 min for xanthine. Increase in adenosine concentration in ischemic relative to control retina-choroid plateaued at 30 min of ischemia, while inosine and hypoxanthine concentrations increased continuously. The increase in xanthine concentration was exponential throughout the measurement period. This study documented the time-related changes in purine nucleoside concentration during ischemia. Prolonged ischemia results in ongoing production of xanthine, which by serving as a precursor for oxygen free radical formation, could be a pathogenic factor in prolonged retinal ischemia.     

Effects of alpha- and beta-adrenoceptor blockade on purine secretion induced by sympathetic nerve stimulation in the rat kidney

To characterize the effects of renal sympathetic nerve activation (RSNA) on renal purine secretion, 13 perfused rat kidneys were stimulated with periarterial electrodes at 7 Hz for 3 min, and purine secretion was determined by measuring with high-performance liquid chromatography purines in the renal venous perfusate 1 min before and during the last minute of RSNA. RSNA significantly increased renal perfusion pressure and significantly increased the secretion of adenosine and adenosine metabolites (inosine, hypoxanthine, and xanthine) by 2- to 5-fold. To investigate the participation of alpha- and beta-adrenoceptors in this response, four groups of perfused kidneys (n = 5/group) were pretreated with either vehicle, prazosin (alpha1-adrenoceptor antagonist; 0.03 microM), phentolamine (alpha1/2-adrenoceptor antagonist; 3 microM), or propranolol (beta1/2-adrenoceptor antagonist; 0.1 microM), and purine secretion was measured before and during RSNA at 1, 3, 5, 7, and 9 Hz. Prazosin, phentolamine, and propranolol abolished the RSNA-induced increase in the secretion of adenosine, inosine, hypoxanthine, and xanthine. In contrast, prazosin and phentolamine nearly abolished, whereas propranolol only slightly reduced, renal vascular responses to RSNA. Our results indicate that RSNA increases renal purine secretion via a mechanism that requires both alpha- and beta-adrenoceptors. It is well known that in the kidney adenosine activates renal afferent nerves, enhances renovascular responses to norepinephrine and angiotensin II, and increases sodium reabsorption; therefore, RSNA-induced adenosine production may contribute to the hypertensive effects of RSNA. Moreover, the antihypertensive effects of beta-adrenoceptor antagonists may in part be due to inhibition of RSNA-induced renal adenosine production.     

Conspiring to succeed: the process of a joint military health care merger

The exact mechanism of hypothermic cerebroprotection after traumatic brain injury (TBI) is not fully understood. The present study was conducted to investigate the effects of mild hypothermia on trauma-induced synthesis of nitric oxide (NO), which has been implicated in the pathogenesis of ischemic brain damage associated with glutamate neurotoxicity. Cerebral contusion was created in the rat parietal cortex by a weight-drop method, and extracellular concentrations of the NO end products nitrite and nitrate were measured using in vivo brain microdialysis and capillary electrophoresis under normothermic (37 degrees C) and mild hypothermic (32 degrees C) conditions. In normothermic animals, the level of NO end products increased markedly 10 min after contusion, reaching a maximum level at 20 min. In the hypothermic rats, such increases were absent. Although it is unknown whether endothelial NO synthase, neuronal NO synthase, or both caused the elevation of the NO end products seen in the normothermic animals, the present results indicate that inhibition of NO synthesis may play a part in hypothermic cerebroprotection following TBI.     

Polyps of the colon and rectum

The uptake and metabolism of [14C]- or E[3H] adenosine have been studied in suspensions of washed platelets and in platelet rich plasma. The appearance of radioactivity in the platelets and the formation of radioactive adenosine metabolites have been used to determine the uptake. Adenosine is transported into human blood platelets by two different systems: a low Km system (9.8 muM) which is competitively inhibited by papaverine, and a high Km system (9.4 mM) which is competitively inhibited by adenine. Adenosine transported via the low Km system is probably directly incorporated into adenine nucleotides, while adenosine transported through the high Km system arrives unchanged inside the platelet and is then converted into inosine and hypoxanthine or incorporated into adenine nucleotides.     

Glycerol as a marker for post-traumatic membrane phospholipid degradation in rat brain

Degradation of membrane phospholipids (PLs) is a well known phenomenon in acute brain injuries and is thought to underlie the disturbance of vital cellular membrane functions. In the present study glycerol, an end product of PL degradation, was examined in brain interstitial fluid as a marker of PL breakdown following experimental traumatic brain injury (TBI) using microdialysis. TBI was induced in artificially ventilated rats using the weight-drop technique. The trauma caused a significant, eight-fold increase of dialysate glycerol in the injured cortex, with the peak concentration in the second 10 min fraction after trauma. Glycerol then levelled off but remained significantly above sham-operated controls for the entire 4 h observation period in the perimeter of the injury region where scattered neuronal death is seen. The results support the concept that PL degradation occurs early after TBI and that interstitial glycerol, harvested by microdialysis, may be useful as a marker allowing in vivo monitoring of PL breakdown.     

Screening a peptidyl database for potential ligands to proteins with side-chain flexibility

OBJECTIVE: Microdialysis and 31P-NMR spectroscopy were used to test opposing hypotheses that ischemic preconditioning inhibits adenine nucleotide degradation and purine efflux, or that preconditioning activates cardiovascular adenosine formation to provide enhanced cardioprotection. METHODS: 31P-NMR spectra and matching interstitial fluid (ISF) or venous effluent samples were obtained from Langendorff perfused rat hearts. Control hearts (n = 9) underwent 30 min of global normothermic ischemia and 30 min reperfusion. Preconditioned hearts (n = 6) were subjected to a 5 min ischemic episode and 10 min reflow prior to 30 min ischemia and 30 min reperfusion. Effects of repetitive ischemia-reperfusion (3 x 5 min ischemic episodes) on adenosine levels and energy metabolism were also assessed (n = 8). RESULTS: Preconditioning improved post-ischemic recovery of heart rate x left ventricular developed pressure (71 +/- 5 vs 43 +/- 8%, P < 0.05) and end-diastolic pressure (14 +/- 3 vs 29 +/- 4 mmHg, P < 0.05) compared with control hearts, respectively. Preconditioning did not alter intracellular ATP, phosphocreatine (PCr), inorganic phosphate (Pi), H+ or free Mg2+ during global ischemia, but improved recoveries of PCr, Pi, and delta GATP on reperfusion. ISF adenosine increased more than 20-fold during 30 min ischemia. The 5 min preconditioning episode increased ISF adenosine 3-fold, and reduced ISF adenosine and inosine during subsequent prolonged ischemia by up to 75%. Venous purine levels during reperfusion were also reduced by preconditioning. Accumulation of adenosine in ISF and venous effluent during repetitive ischemia was progressively reduced despite comparable changes in substrate for adenosine formation via 5'-nucleotidase, (5'-AMP), and in allosteric modulators of this enzyme (Mg2+, H+, Pi, ADP, ATP). CONCLUSIONS: (i) Ischemic preconditioning reduces interstitial and vascular adenosine levels during ischemia-reperfusion, (ii) reduced ISF adenosine during ischemia is not due to reduced ischemic depletion of adenine nucleotides in preconditioned rat hearts, (iii) preconditioning may inhibit adenosine formation via 5'-nucleotidase in ischemic rat hearts, and (iv) improved functional recovery with preconditioning is unrelated to metabolic/bioenergetic changes during the ischemic insult, but may be related to improved post-ischemic recovery of [Pi] and delta GATP in this model.     

Fumonisin B1 contamination of maize and experimental acute fumonisin toxicosis in pigs

As part of the stress response, the 72 kDa heat shock protein (hsp72) is induced in neurons after ischemic and traumatic brain injury (TBI). To examine the stress response after TBI with secondary insult, we examined the regional and cellular expression of hsp72 mRNA and protein after controlled cortical impact (CCI) injury with secondary hypoxemia and mild hypotension in rats. Rats were killed at 6, 8, 24, 72, or 168 h after trauma. Naive and sham-operated rats were used as controls. Brains were removed, and in situ hybridization (n = 2/group), immunocytochemistry (n = 4/group), and Western blot analysis (n = 3 to 5/group) for hsp72 was performed. Hsp72 mRNA was expressed in neurons in the ipsilateral cortex, CA3 region of the hippocampus, hilus, and dentate gyrus at 6 h. Hsp72 mRNA was expressed primarily in the ipsilateral cortex, at 24 h, and by 72 h hsp72 mRNA expression returned to near basal levels. Hsp72 protein was seen in ipsilateral cortical neurons, hilar neurons, and neurons in the medial aspect of the CA3 region of the hippocampus (CA3-c) at 24 h. At 72 h, hsp72 immunoreactivity was reduced versus 24 h in these same regions, but it was increased versus baseline. Western blot analysis confirmed an increase in hsp72 protein in the ipsilateral cortex. The regional pattern of hsp72 mRNA induction in neurons was similar to the pattern of protein expression after CCI, with the exceptions that hsp72 mRNA, but not protein, was expressed in the dentate gyrus and the lateral aspect of the CA3 region of the hippocampus (CA3-a). The stress response, as detected by hsp72 expression, is induced in some neurons in some regions that are selectively vulnerable to delayed neuronal death in this model of TBI. The failure to translate some proteins including hsp72 may be associated with delayed neuronal death in certain hippocampal regions after TBI.     

Autoxidation of pyridoxalated hemoglobin polyoxyethylene conjugate

Vascular endothelial growth factor (VEGF) is an endothelial cell mitogen which stimulates angiogenesis. VEGF is regulated by multiple factors such as hypoxia, phorbol esters, and growth factors. However, data concerning the expression of VEGF in the different vascular cell types and its regulation by cAMP are not available. In the present study, we have investigated the effect of adenylate cyclase activation on VEGF mRNA expression in rat vascular cells in primary culture. Basal VEGF expression is greater in smooth muscle cells than in endothelial cells and fibroblasts. A 4-h treatment with forskolin (10(-5) M) induced a 2-fold stimulation of VEGF mRNA expression in smooth muscle cells and fibroblasts, but, in contrast, did not affect VEGF expression in endothelial cells. In smooth muscle cells, a pharmacologically induced increase in intracellular cAMP levels using iloprost or isoprenaline led to a rise in VEGF mRNA expression comparable to that induced by forskolin. Adenosine, which increases cAMP levels in smooth muscle cells, also increases VEGF expression. Moreover, the 2.2-fold stimulation of VEGF expression by adenosine was enhanced following a cotreatment with cobalt chloride (a hypoxia miming agent). The observed additive effect (4.3-fold increase) suggests that these two factors, hypoxia and adenosine, regulate VEGF mRNA expression in smooth muscle cells by independent mechanisms.     

The influence of the milieu on the rate of neutralisation of herpes simplex virus 1

Minimizing secondary injury after severe traumatic brain injury (TBI) is the primary goal of cerebral resuscitation. For more than two decades, hyperventilation has been one of the most often used strategies in the management of TBI. Laboratory and clinical studies, however, have verified a post-TBI state of reduced cerebral perfusion that may increase the brain's vulnerability to secondary injury. In addition, it has been suggested in a clinical study that hyperventilation may worsen outcome after TBI. OBJECT: Using the controlled cortical impact model in rats, the authors tested the hypothesis that aggressive hyperventilation applied immediately after TBI would worsen functional outcome, expand the contusion, and promote neuronal death in selectively vulnerable hippocampal neurons. METHODS: Twenty-six intubated, mechanically ventilated, isoflurane-anesthetized male Sprague-Dawley rats were subjected to controlled cortical impact (4 m/second, 2.5-mm depth of deformation) and randomized after 10 minutes to either hyperventilation (PaCO2 = 20.3 +/- 0.7 mm Hg) or normal ventilation groups (PaCO2 = 34.9 +/- 0.3 mm Hg) containing 13 rats apiece and were treated for 5 hours. Beam balance and Morris water maze (MWM) performance latencies were measured in eight rats from each group on Days 1 to 5 and 7 to 11, respectively, after controlled cortical impact. The rats were killed at 14 days postinjury, and serial coronal sections of their brains were studied for contusion volume and hippocampal neuron counting (CA1, CA3) by an observer who was blinded to their treatment group. Mortality rates were similar in both groups (two of 13 in the normal ventilation compared with three of 13 in the hyperventilation group, not significant [NS]). There were no differences between the groups in mean arterial blood pressure, brain temperature, and serum glucose concentration. There were no differences between groups in performance latencies for both beam balance and MWM or contusion volume (27.8 +/- 5.1 mm3 compared with 27.8 +/- 3.3 mm3, NS) in the normal ventilation compared with the hyperventilation groups, respectively. In brain sections cut from the center of the contusion, hippocampal neuronal survival in the CA1 region was similar in both groups; however, hyperventilation reduced the number of surviving hippocampal CA3 neurons (29.7 cells/hpf, range 24.2-31.7 in the normal ventilation group compared with 19.9 cells/hpf, range 17-23.7 in the hyperventilation group [25th-75th percentiles]; *p < 0.05, Mann-Whitney rank-sum test). CONCLUSIONS: Aggressive hyperventilation early after TBI augments CA3 hippocampal neuronal death; however, it did not impair functional outcome or expand the contusion. These data indicate that CA3 hippocampal neurons are selectively vulnerable to the effects of hyperventilation after TBI. Further studies delineating the mechanisms underlying these effects are needed, because the injudicious application of hyperventilation early after TBI may contribute to secondary neuronal injury.     

Development of a silicon-mediated [3 + 2] annulation and its application to the synthesis of natural products]

Hearts preconditioned by brief ischemia are characterized by a reduced rate of cellular purine metabolite production during subsequent prolonged ischemia; the purpose of this study was to determine if transient exogenous adenosine pretreatment can mimic this phenomenon. The accumulation of interstitial fluid (ISF) purine metabolites during prolonged ischemia in untreated anesthetized dogs (n = 7) was compared to that in a group pretreated with brief ischemia (ischemic preconditioned group; n = 9), a group pretreated with 1.5 micromoles/min intracoronary adenosine (n = 7), and a group pretreated with 100 micromoles/min intracoronary adenosine (n = 7). Ischemic preconditioning was achieved by a 5 min period of left anterior descending coronary artery (LAD) occlusion followed by 10 min of reperfusion. The adenosine-treated groups were subjected to 10 min of intracoronary adenosine followed by 10 min of recovery. All animals were exposed to 60 min LAD occlusion followed by 60 min reperfusion. The changes in ISF adenosine and adenosine metabolites were assessed by cardiac microdialysis, using dialysate concentrations as indices of ISF levels. Ischemic preconditioning decreased the rate of dialysate adenosine and total purine accumulation during the prolonged ischemia. Although the two doses of exogenous adenosine bracketed the increase in ISF adenosine seen with ischemic preconditioning, neither adenosine dose was able to attenuate the rate of purine metabolite accumulation during prolonged ischemia. We conclude that exogenous adenosine pretreatment is unable to mimic the reduced ischemia-induced purine efflux that is characteristic of myocytes pretreated with brief ischemia.     

Combined therapy affects outcomes differentially after mild traumatic brain injury and secondary forebrain ischemia in rats

Muscarinic and NMDA receptors contribute to post-traumatic hypersensitivity to secondary ischemia. However, the effect of these receptor antagonists on behavior and CA1 neuronal death after traumatic brain injury (TBI) with acute (1 h after TBI) forebrain ischemia has not been systematically assessed. We examined cognitive and motor dysfunction and the relationship of behavior deficits to neuronal death in this model using muscarinic and NMDA antagonists. Three behavioral groups (n=10/group) of Wistar rats were subjected to mild TBI and 6 min of forebrain ischemia imposed 1 h after TBI with 45 days survival. Motor and spatial memory performance were assessed using the rotarod task and Morris water maze. Seven additional groups (n=6/group) were evaluated only for CA1 death after 7 days survival following sham, individual or combined injury with and without drug treatments. Rats were given 0.3 mg/kg MK-801 (M) and 1.0 mg/kg scopolamine (S) alone or combined (M-S) before or 45 min after TBI. Rotarod performance was tested at days 1-5 and maze performance on days 11-15 and 40-44 after M-S treatment. The 7-day studies showed M-S treatment (p<0.01) reduced CA1 neuronal death better than either S or M alone. Behavioral groups had inadvertent post-ischemic hypothermia that decreased CA1 death and likely influenced behavioral morbidity. M-S given before TBI (p<0.01) decreased memory deficits on day 15, while M-S treatment given after TBI was ineffective. Unexpectedly, M-S treatment before or after TBI produced transient motor deficits (p<0. 01). Memory improvement occurred independent of CA1 death.     

Gastroduodenal lesions and non-steroidal anti-inflammatory agents. What role does Helicobacter pylori play in this relationship?

In order to elucidate the mechanisms of release of EAAs and their excitotoxicity in cerebral contusion, cortical contusion was produced in the rat parietal cortex, and the changes in extracellular levels of EAAs in the central and peripheral areas of contusion were investigated using microdialysis. The cortical contusion induced a rapid increase in dialysate concentration of glutamate ([Glu]d) from a baseline level of 4.6+/-2.8 microM to a maximum level of 36.3+/-12.8 microM. This elevation of glutamate was significantly attenuated by mild hypothermia (32 degrees C for 90 min, comprising 20 min before and 70 min after the injury induction) in the peripheral area of contusion (p < 0.01) but not in the central area. Histological evaluations revealed that the hypothermia reduced the necrosis volume of contusion to 38.3% of that in the normothermic control (p < 0.01). In situ administration of Co2+, an inhibitor of Co2+-dependent exocytotic release of EAAs from the nerve terminals, via the microdialysis system, also attenuated the [Glu]d elevation following cortical contusion, in the peripheral area of contusion (p < 0.01) but not in the central area. These findings indicate that cerebral contusion involves heterogeneous and complex mechanisms of EAA release into the extracellular space. The release of EAAs in the contusion core was nonsensitive to hypothermia and Co2+ administration, suggesting that such EAA release was related to primary disruption of the cell membrane or vascular wall by the physical force of the head trauma, resulting in leakage of EAAs from the metabolic pool in the cytosole or blood stream. In contrast, in the peripheral area, the effectiveness of hypothermia and Co2+ administration implied a presynaptic mechanism of EAA release, which consisted, at least in part, of Ca2+-dependent exocytotic EAA release from depolarized nerve terminals. The EAAs released in the contusion core may diffuse towards a peripheral direction and act on the postsynaptic receptors, causing neuronal depolarization. Such a diffusion-reaction process appears to induce additional release of EAAs from the depolarized nerve terminals. Hypothermia may block this diffusion-reaction process and eventually reduce the contusion volume.     

Intrathecal adenosine administration: a phase 1 clinical safety study in healthy volunteers, with additional evaluation of its influence on sensory thresholds and experimental pain

BACKGROUND: Several animal studies show antinociceptive effects of intrathecally administered adenosine and its analogs. However, there is no clinical experience regarding the effects of intrathecal adenosine in humans. METHODS: The side effects and analgesic effects of intrathecal adenosine (500-2,000 microg) on experimental pain were studied in 12 healthy volunteers. Before and after adenosine was given, the authors evaluated the cold pain rating of the foot (submersion in ice water for 1 min), the forearm ischemic pain rating during a 30-min tourniquet test, and the thermal and tactile pain thresholds on healthy and inflamed skin after application of mustard oil (4 min) to the calf. The areas of secondary allodynia surrounding the inflammation were also determined. The cerebrospinal fluid level of adenosine was determined before and after injection. RESULTS: Intrathecal adenosine caused a 1,000- to 2,000-fold elevation of the cerebrospinal fluid concentration. One volunteer experienced transient (30 min) lumbar pain after injection at a dose of 2,000 microg. There were no other complications in any other volunteers. Adenosine reduced, in a non-dose-dependent manner, the areas of secondary allodynia after skin inflammation (brush, P < 0.06; and von Frey hair, P < 0.03) and reduced the forearm tourniquet ischemic pain rating (P = 0.01). Tactile pain thresholds were significantly reduced by mustard oil inflammation during control, whereas adenosine treatment prevented this reduction. The ice water-induced cold pain rating was not influenced by adenosine. CONCLUSIONS: An intrathecal adenosine injection of 1,000 microg lacked side effects in healthy volunteers. The compound attenuated different types of experimental pain.     

Eosinophilic myopathic syndromes

The purpose of this study was to develop a standardised maximal treadmill exercise test performed until fatigue in order to find reproducible markers for anaerobic metabolism, specifically adenine nucleotide degradation. Six Standardbred trotters performed an incremental maximal treadmill exercise test in 1 min steps (starting with 7 m/s) until they could no longer keep pace with the treadmill. The test was performed twice with at least one week between the tests. Heart rate was recorded and venous blood samples were obtained during the test and in the recovery period for determination of plasma lactate, hypoxanthine, xanthine and uric acid. Muscle biopsy samples (m. gluteus) were collected at rest, immediately post exercise, and after 15 min recovery and analysed for their concentrations of glycogen, creatine phosphate (CP), adenosine triphosphate (ATP), adenosine diphosphate (ADP), adenosine monophosphate (AMP), inosine monophosphate (IMP) and muscle lactate (MLa). Significant decreases in glycogen, CP and ATP and significant increases in IMP and MLa were seen immediately post exercise. None of these metabolites had returned to resting levels after 15 min of recovery. A marked increase in plasma lactate (PLa) occurred during exercise and the peak concentration (mean value = 27.2 mmol/l) was reached within 5 min of recovery. Plasma uric acid concentration did not increase during exercise but rose markedly immediately post exercise, reaching the highest level (mean value = 121.5 micromol/l) at 20-30 min recovery. The duration of the maximal test was related to peak PLa and the uric acid concentration at 30 min of recovery. A correlation was also found between the ATP and IMP concentrations immediately post exercise and the plasma uric acid concentration at 30 min of recovery. The results show that this treadmill test triggered anaerobic metabolism and also that uric acid concentration post exercise seems to be a marker for the adenine nucleotide degradation that occurs during intense exercise. No significant differences were seen in metabolic response between the 2 test occasions.     

Modulation of adenosine effects in attenuation of ischemia and reperfusion injury in rat heart

We investigated whether xanthine oxidase-derived superoxide radical generation could be modified by interfering with adenosine transport and metabolism in reducing myocardial injury during post-ischemic reperfusion. Isolated rat hearts perfused at constant pressure were subjected to 20 min of pretreatment with test agents, followed by 40 min global ischemia and 30 min reperfusion with or without test agents. In hearts treated with adenosine deaminase inhibitor, erythro 9-(2-hydroxy-3-nonyl) adenine (EHNA), alone or together with a selective nucleoside transport blocker, p-nitrobenzylthioinosine (NBMPR), the accumulated amount of O-2. was significantly reduced [10.2+/-0.97, 11.6+/-2.4, 8.1+/-0.51, respectively, v 31.6+/-2.1 (s. e.) nmol/wet g/30 min in ischemic control, P<0.01]. A positive correlation between O-2. and inosine release was observed in the initial 5 min of reperfusion in hearts treated with either EHNA or NBMPR ( r=0.475, P<0.05). Furthermore, the accumulated amount of LDH release showed positive correlation with that of O-2. among the same groups (r=0.474, P<0.05). Both EHNA and NBMPR had the cardioprotective effect on the recovery of left ventricular end-diastolic pressure (LVEDP), ATP repletion, and build up of endogenous adenosine. This study suggests that : (1) adenosine metabolism can be manipulated towards the formation of O-2. during reperfusion, and it has an important bearing on the cardiac recovery of ischemic myocardium, (2) the generation of O-2. is related to only inosine release during initial reperfusion.     

Interpersonal Stressors and Resources as Predictors of Parental Adaptation Following Pediatric Traumatic Injury.

The authors examined the relationship of preinjury interpersonal resources and stressors to parental adaptation following pediatric traumatic brain injury (TBI) and orthopedic injury. Parents of children with severe TBI (n = 53), moderate TBI (n = 56), and orthopedic injuries (n = 80) were assessed soon after injury, 6 and 12 months after the initial evaluation, and at an extended follow-up with a mean of 4 years postinjury. General linear model analyses provide support for both main and moderating effects of stressors and resources on parental adjustment. Support from friends and spouse was associated with less psychological distress, whereas family and spouse stressors were associated with greater distress. The results also reveal a marked decline in injury-related stress over follow-up for families in the severe TBI group who reported a combination of high stressors and high resources. The decline suggests that interpersonal resources attenuated long-term family burden because of severe TBI. The findings are discussed in terms of their implications for intervention following TBI. (PsycINFO Database Record (c) 2010 APA, all rights reserved)     

Adenosine transport inhibition ameliorates postischemic hypoperfusion in pigs

Cerebral ischemia is often followed by a period of delayed hypoperfusion that may contribute to tissue injury. We tested the hypothesis that augmentation of interstitial adenosine can improve tissue perfusion under this condition 10 min global ischemia was produced in two groups of isoflurane-anesthetized newborn pigs by occlusion of subclavian and brachiocephalic arteries, and changes in local cortical blood flow and cortical interstitial purine metabolites were measured using the combined hydrogen clearance-microdialysis technique. In one group, the dialysis probe was perfused with artificial cerebrospinal fluid buffer containing nitrobenzyl-thioinosine (NBT1, 100 mumol/l), a competitive inhibitor of adenosine transport. In the untreated group (n = 9), baseline cortical blood flow (39 +/- 3 ml/min/100 g) was depressed by 51 +/- 5% and 42 +/- 6% at 40 and 60 min, respectively, of postischemic reperfusion. NBTI increased baseline interstitial adenosine levels 2.4-fold which increased baseline cortical blood flow 1.5-fold to 60 +/- 4 ml/min/100 g, and increased both absolute adenosine levels as well as adenosine as a percentage of total purine metabolites throughout ischemia and reperfusion. As a result, the extent of postischemic hypoperfusion was significantly lessened in NBTI-treated animals (n = 9), with reductions in cortical blood flow of only 28 +/- 3% and 24 +/- 5% at 40 and 60 min of reperfusion, respectively. These results indicate that inhibition of adenosine transport by NBTI elevates interstitial adenosine concentration during and following cerebral ischemia, and concomitantly improves cortical perfusion in the post-ischemic period. The latter effect may contribute to the documented neuroprotective efficacy of adenosinergic therapy in cerebral ischemia.