Influence of morphine on medial prefrontal cortex alpha2 adrenergic system in passive avoidance learning in rats.

The prelimbic region of the medial prefrontal cortex (mPFC) is a brain area crucial for memory, attention, and decision making. It has been shown that α2-adreneoceptors (α2-ARs) play a powerful role in regulating memory and attention functions in this region. Since many studies have demonstrated the impairment effect of morphine on memory through mPFC, we aimed to investigate the possible interaction between α2-ARs of the mPFC and morphine induced amnesia in passive avoidance learning in rats. Animals were bilaterally implanted with chronic cannulas in the mPFC, trained in the step-through type passive avoidance task, and tested 24h after training; step-through latencies were measured. Our data indicate that post-training i.p. administration of morphine (2.5, 5 and 7.5mg/kg) dose-dependently reduced the step-through latency, showing an amnesic effect. Post-training intra-mPFC administration of yohimbine (an α2-adrenergic antagonist, 0.125, 0.25 and 0.5µg/rat) and clonidine (an α2-adrenergic agonist, 0.001, 0.01 and 0.2µg/rat), dose dependently impaired memory retrieval. Furthermore, post-training intra-mPFC microinjection of ineffective doses of yohimbine or clonidine significantly reversed the inhibitory effect of morphine on memory retrieval. Furthermore, SKF96365 (a presynaptic calcium channel blocker) reduced yohimbine and showed slight inhibition of clonidine effect. These results suggest that α2-ARs of the mPFC may play an important role in morphine-induced amnesia.

Tramadol antinociception is potentiated by clonidine through α2-adrenergic and I2-imidazoline but not by endothelin ET(A) receptors in mice.

Tramadol is a centrally acting analgesic that acts via µ-opioid agonism and by blocking the neuronal uptake of norepinephrine and serotonin. Clonidine potentiates the antinociceptive effects of tramadol; however the receptors involved in this potentiation have not been studied. Endothelin ET(A) receptor antagonists potentiate antinociceptive effects of morphine and oxycodone; however the effects of endothelin ET(A) receptor antagonists on tramadol antinociception have not been evaluated. This study was conducted to determine the effect of clonidine on tramadol antinociception; the role of opioid, α2-adrenergic and I2-imidazoline receptors in clonidine potentiation of tramadol antinociception; and the effect of endothelin ET(A) receptor antagonists in modulating tramadol antinociception. Antinociceptive (tail-flick and hot-plate) latencies were measured in male Swiss Webster mice treated with tramadol; clonidine plus tramadol; or antagonists plus tramadol. Mice were pretreated with naloxone (opioid antagonist), yohimbine (α2-adrenoceptor antagonist), idazoxan (α2-adrenoceptor/I2-imidazoline antagonist), BMS182874 or BQ123 (endothelin ET(A) receptor antagonists) to study the involvement of these receptors. Tramadol produced a dose dependent increase in antinociceptive latencies. Tramadol antinociception was partially blocked by naloxone but not by yohimbine or idazoxan. Clonidine potentiated tramadol antinociception; potentiation was blocked by naloxone, yohimbine and idazoxan. Idazoxan produced a more pronounced blockade of potentiation than yohimbine. BMS182874 or BQ123 had no effect on tramadol antinociception, indicating that endothelin ET(A) receptors are not involved in tramadol antinociception in mice. Results demonstrate the involvement of opioid but not α2-adrenergic/I2-imidazoline receptors in tramadol antinociception and that opioid, α2-adrenergic and I2-imidazoline receptors are involved in clonidine potentiation of tramadol antinociception.

α(2) Adrenergic and imidazoline receptor agonists prevent cue-induced cocaine seeking.

BACKGROUND: Drug-associated cues can elicit stress-like responses in addicted individuals, indicating that cue- and stress-induced drug relapse may share some neural mechanisms. It is unknown whether α(2) adrenergic receptor agonists, which are known to attenuate stress-induced reinstatement of drug seeking in rats, also reduce cue-induced reinstatement. METHODS: Rats were tested for reinstatement of drug seeking following cocaine self-administration and extinction. We first evaluated the effects of clonidine, an agonist at α(2) and imidazoline-1 (I(1)) receptors, on relapse to cocaine seeking. To explore possible mechanisms of clonidine's effects, we then tested more specific α(2) or I(1) agonists, postsynaptic adrenergic receptor (α(1) and ß) antagonists, and corticotropin-releasing factor receptor-1 antagonists. RESULTS: We found that clonidine, and the more selective α(2) agonists UK-14,304 and guanfacine, decreased cue-induced reinstatement of cocaine seeking. The specific I(1) receptor agonist moxonidine reduced cue-induced as well as cocaine-induced reinstatement. Clonidine or moxonidine effects on cue-induced reinstatement were reversed by the selective α(2) receptor antagonist RS-79948, indicating a role for α(2) receptors. Prazosin and propranolol, antagonists at the α(1) and ß receptor, respectively, reduced cue-induced reinstatement only when administered in combination. Finally, the corticotropin-releasing factor receptor-1 antagonist CP-154,526 reduced cue-induced reinstatement, as previously observed for stress-induced reinstatement, indicating possible overlap between stress and cue mechanisms. CONCLUSIONS: These results indicate that α(2) and I(1) receptor agonists are novel therapeutic options for prevention of cue-induced cocaine relapse. Given that α(2) receptor stimulation is associated with sedation in humans, the I(1) agonist moxonidine seems to have substantial potential for treating addictive disorders.

Clonidine in horses: identification, detection, and clinical pharmacology.

Clonidine is classified as a class 3 performance-enhancing agent by the Association of Racing Commissioners International and thus has the potential to influence the outcome of a race. In this study, the authors developed and validated a sensitive gas chromatograph and mass spectrometer method to determine the pharmacokinetic parameters of clonidine in equine plasma samples after IV administration of a single dose (0.025 mg/kg) of clonidine in horses. At this dose, clonidine produced rapid and profound sedation, which cold be quickly reversed with yohimbine. Clonidine was able to produce an analgesic effect but failed to provide maximal analgesia in all horses; the limited analgesic effect persisted for about 60 minutes.

Chloroethylclonidine reveals that alpha (1 A)-adrenoceptors mediate contraction in aorta of alpha (1 D)-adrenoceptor knockout mice.

1 We have characterized the alpha(1)-adrenoceptor subtypes present in isolated aorta of the alpha(1D)-adrenoceptor knockout (KO) mice, by chloroethylclonidine (CEC)-induced alkylation and their protection by selective alpha(1)-adrenoceptor antagonists. 2 The alpha(1D)-adrenoceptor is involved in the contractile response to noradrenaline in wild type (WT) mouse aorta. 3 In WT mice 5-methylurapidil (5-MU, an alpha(1A)-adrenoceptor antagonist) or BMY 7378 (8-[2-[4-(2-methoxyphenyl)-1-piperazinyl] ethyl]-8-azaspiro[4.5] decane-7,9 dione, a selective alpha(1D)-adrenoceptor antagonist), protected the receptors from CEC-induced (alpha(1B/D)-adrenoceptor) alkylation, the combination of both antagonists resulted in complete protection, while AH11110A (1-[biphenyl-2-yloxy]-4-imino-4-piperidin-1-yl-butan-2-ol, an alpha(1B)-adrenoceptor antagonist) did not protect. 4 In aorta of KO mice there was a 19-fold rightward shift in noradrenaline effective concentration (EC(50)) compared with WT; while 5-MU alone or in combination with AH11110A protected alpha(1)-adrenoceptors to the same extent. 5 The data indicate that alpha(1A)-adrenoceptors mediate contraction and suggest their role in maintaining homeostasis in the alpha(1D)-adrenoceptors KO mice.

Effects of alpha- and beta-adrenergic agonists on Toxoplasma gondii infection in murine macrophages.

We investigated the effects of alpha- and beta-adrenergic receptor agonists on the ability of Toxoplasma gondii to infect and proliferate in cultured murine macrophages. Macrophages pretreated in vitro with varying concentrations of alpha- and beta-adrenergic agonists and incubated with the RH strain of T. gondii did not result in a significant increase in the percentage of infected macrophages compared with negative controls. When parasites were pretreated with L-phenylephrine, an alpha-agonist, and L-isoproterenol, a beta-agonist, before infection, there was no significant change in the percentage of infected macrophages. Clonidine, an alpha2-adrenergic agonist, led to a significant decrease in the number of infected macrophages at all concentrations tested. The effects of clonidine were blocked by yohimbine, a specific alpha2-adrenergic antagonist, but not by phentolamine, an alpha1-adrenergic antagonist. These results suggest that the antiparasitic effects exhibited by clonidine (alpha2-adrenergic agonist) are mediated through an alpha2-adrenoreceptor found on the surface of T. gondii.

Intrathecal neostigmine prevents intrathecal clonidine from attenuating hypercapnic cerebral vasodilation in rabbits.

We previously demonstrated that lumbar intrathecal alpha(2) agonists attenuate hypercapnia-induced cerebral vasodilation. The combination of intrathecal clonidine and neostigmine is being investigated as pain therapy. The effects of their combination on cerebrovascular reactivity are unknown. We allocated rabbits anesthetized with pentobarbital to two groups: (a) clonidine (normal saline followed 30 min later by clonidine 2 microg/kg, both into the lumbar intrathecal space; n = 6), and (b) neostigmine-pretreatment (neostigmine 2 microg/kg followed 30 min later by clonidine 2 microg/kg, both into the lumbar intrathecal space; n = 6). We then evaluated the hypercapnia-induced changes in pial arteriolar diameter in these two groups using the closed cranial window preparation. The pial arteriolar dilator response to hypercapnia was significantly attenuated in the clonidine group (14% +/- 4%, 4% +/- 4%, 6% +/- 6%, and 5% +/- 7% for before and 30, 60, and 90 min, respectively). Neither normal saline nor neostigmine alone induced any change in the cerebral reactivity to hypercapnia. Pretreatment with neostigmine completely prevented the clonidine-induced attenuation of the hypercapnic cerebral vasodilation attenuated by intrathecal clonidine (16% +/- 7%, 15% +/- 6%, 12% +/- 6%, and 16% +/- 8%, respectively).

Mechanism of the endothelium-dependent vasodilation and the antihypertensive effect of Brazilian red wine.

The mechanisms involved in the cardioprotector effect of red wine have not yet been completely elucidated but probably an endothelium-dependent vasodilator action may play a significant role in this effect. Experiments were undertaken to determine whether a Brazilian red wine (BRW) induces vasodilation in the mesenteric vascular bed (MVB) and an antihypertensive effect was also assessed in rats with NO-deficient hypertension. In MVB precontracted with norepinephrine, BRW (alcohol-free lyophilized) induces a long-lasting endothelium-dependent vasodilation that is not reduced by indomethacin. Inhibition of NO-synthase by NG-nitro-L-arginine methyl ester (L-NAME) and guanylyl cyclase by 1H-[1,2,3] oxadiazolo [4,4-a]quinoxalin-1-one (ODQ) reduces the vasodilator effect of BRW. In vessels precontracted with norepinephrine and depolarized with KCl (25 Mm) or treated with Ca-dependent K channel blockers charybdotoxin (ChTx) plus apamin, the effect of BRW was significantly reduced. However, this effect is not affected by ATP-dependent K (KATP) channel blocker (glibenclamide). The residual vasodilator effect of BRW observed in vessels pretreated with ChTx plus apamin is completely abolished by ChTx plus apamin plus L-NAME. Concentrations of atropine, pyrilamine, yohimbine, and HOE 140 that significantly reduced the vasodilator effect of acetylcholine, histamine, clonidine, and bradykinin, respectively did not change the vasodilator effect of BRW. Chronic oral administration of BRW induced a significant reduction in systolic, mean and diastolic arterial pressure in rats with L-NAME hypertension. The present results demonstrated that vasodilator effect of BRW is dependent on endothelium-derived hyperpolarizing factor (EDHF) in combination with nitric oxide (NO). The antihypertensive effect of red wine demonstrated in the present study may play a significant role on the cardioprotective action of chronic red wine consumption.

Clonidine induces rat aorta relaxation by nitric oxide-dependent and -independent mechanisms.

The alpha1- and alpha2-adrenoceptors coexist in vascular smooth muscle cells producing vascular contraction and relaxation. This study was designed to investigate which is the mechanism activated by clonidine in the rat aorta, and the endothelial factors possibly involved in the relaxation induced by clonidine. The alpha2-adrenoceptors agonist clonidine relaxed rat aortas pre-contracted with phenylephrine, with or without endothelium. In non-contracted denuded arteries, clonidine produced contractions instead of relaxation. In intact endothelium aortic rings, clonidine induced greater relaxation than in denuded aortic rings. In aortas with intact endothelium, the NO-synthase inhibitor L-NAME (10 micromol/L) and the NO-scavenger hemoglobin (10 micromol/L) reduced the relaxation to clonidine. On the other hand, indomethacin (10 micromol/L) failed to alter the relaxation induced by clonidine. These results suggest the participation of NO, but not prostacyclin in clonidine-induced relaxation. In aortic rings pre-contracted with KCl (60 mmol/L) the relaxation induced by clonidine was abolished; however, the K+ channel blockers glibenclamide (K(ATP)), tetraethylamonium (K(Ca)), and the combination of apamin and charybdotoxin (K(Ca)) did not change the relaxation induced by clonidine. The relaxation induced by clonidine on PGF2alpha-contracted arteries was not affected by prazosin. However, in the absence of prazosin, clonidine had an additional contractile effect in PGF2alpha-contracted arteries. In conclusion, our results show that in rat aorta clonidine can activate alpha2-adrenoceptors in the smooth muscle cells and alpha2-adrenoceptors in the endothelial cells that activates NO production, but not prostacyclin and/or EDHF. In the absence of phenylephrine and prazosin, clonidine can also activate alpha1-adrenoceptors and rat aorta contraction.

Blockade of N-methyl-D-aspartate receptors within the rostral ventrolateral medulla antagonizes clonidine-induced cardiovascular effects.

There is wide agreement that the rostral ventrolateral medulla (RVLM) plays a crucial role in the regulation of blood pressure (BP), and that there may be a close correlation between the actions of centrally acting antihypertensive agents and N-methyl-D-aspartate (NMDA) receptor functional states. The present study was done to test the hypothesis that NMDA receptors within the RVLM were involved in the cardiovascular effects of centrally acting antihypertensive drug clonidine in anesthetized and paralyzed rats. Prior unilateral microinjection of NMDA receptor antagonist dizocilpine (MK801, 500 pmol) into the RVLM significantly attenuated (p<0.01, n=9) the reductions of BP (-24+/-6 to -8+/-4 mm Hg) and heart rate (-49+/-9 to -14+/-7 bpm) induced by unilaterally injected clonidine (5 nmol) into the RVLM. Prior bilateral microinjection of MK801 (500 pmol for each side) into the RVLM effectively (p<0.01, n=7) antagonized the hypotension (-25+/-5 to -8+/-2 mm Hg) and bradycardia (-43+/-7 to -11+/-4 bpm) of intravenously administered clonidine (10 microg kg(-1)). Importantly, iontophoretic application of MK801 (60 nA) significantly (p<0.01, n=9) prevented the inhibitory effect of intravenously (10 microg kg(-1)) injected clonidine on the discharge of presympathetic neurons in the RVLM (neuronal inhibition: -39+/-6 to -10+/-2%). In conclusion, the present study shows that the RVLM administrated MK801 effectively antagonizes clonidine-induced cardiovascular effects, and suggests that NMDA receptors within the RVLM contribute to clonidine actions.

Effect of ethanol on reductions in norepinephrine electrochemical signal in the rostral ventrolateral medulla and hypotension elicited by I1-receptor activation in spontaneously hypertensive rats.

BACKGROUND: The mechanism of the antagonistic hemodynamic interaction between ethanol and centrally acting sympatholytics is not known. In this study, we tested the hypothesis that the imidazoline (I1)-receptor modulation of norepinephrine (NE) release within the rostral ventrolateral medulla (RVLM) plays a pivotal role in this clinically relevant hemodynamic interaction. METHOD In anesthetized spontaneously hypertensive rats, the effects of centrally acting sympatholytics on RVLM NE electrochemical signal were investigated by in vivo electrochemistry along with cardiovascular responses in the absence and presence of ethanol. In vivo microdialysis in conscious spontaneously hypertensive rats was used to confirm the electrochemical findings. RESULTS Clonidine (30 microg/kg, intravenously) or rilmenidine (400, 600, or 800 microg/kg) significantly reduced RVLM NE electrochemical signal (index of neuronal activity) and mean arterial pressure; rilmenidine effects were dose-related, and ethanol (1 g/kg) counteracted these responses. Ethanol (1 g/kg) pretreatment increased both RVLM NE electrochemical signal and blood pressure but did not influence the reductions in both variables elicited by subsequently administered clonidine. The alpha2-adrenergic antagonist 2-methoxyidazoxan (30 microg/kg) counteracted rilmenidine (800 microg/kg)-evoked responses. In vivo microdialysis in conscious spontaneously hypertensive rats confirmed the electrochemical findings since clonidine- (30 microg/kg, intravenously) evoked reductions in RVLM NE and the associated hypotension were counteracted by ethanol (1 g/kg). CONCLUSIONS (1) Ethanol counteracts centrally mediated hypotension, at least in part, by increasing RVLM NE; (2) the interaction involves the I1 receptor modulation of RVLM neuronal activity; (3) the alpha2-adrenergic receptor contributes to the electrochemical and cardiovascular effects of high doses of rilmenidine, and (4) the RVLM is a neuroanatomical target for systemically administered ethanol.

Clonidine-induced coronary vasodilatation in isolated guinea pig heart is not mediated by endothelial alpha2 adrenoceptors.

Functional role of endothelial alpha(2)-adrenoceptor in coronary circulation remains unclear. Clonidine, an agonist of alpha(2)-adrenoceptors, was reported to induce coronary vasodilatation via stimulation of endothelial alpha(2)-adrenoceptors or coronary vasoconstriction involving vascular smooth muscle alpha(2)-adrenoceptors. Moreover, H(2) receptor-dependent responses to clonidine were described. Here, we reassess the contribution of endothelial alpha(2)-adrenoceptor and H(2) receptors to coronary flow and contractility responses induced by clonidine in the isolated guinea pig heart. We found that clonidine (10(-9) - 10(-6) M) produced concentration-dependent coronary vasoconstriction without a significant change in contractility. This response was inhibited by the alpha(1)/alpha(2)-adrenoceptor antagonist - phentolamine (10(-5) M) and the selective alpha(2)-adrenoceptor antagonist yohimbine (10(-6) M), but it was not changed by the selective alpha(1)-adrenoceptor antagonist prazosin (10(-6) M). In the presence of nitric oxide synthase inhibitor, L-NAME (10(-4) M) the clonidine-induced vasoconstriction was potentiated. Clonidine at high concentrations of 10(-5) - 3 x 10(-5) M produced coronary vasodilatation, and an increase in myocardial contractility. These responses were abolished by a selective H(2)-receptor antagonist, ranitidine (10(-5) M), but not by phentolamine (10(-5) M). We conclude that in the isolated guinea pig heart, clonidine-induced vasoconstriction is mediated by activation of smooth muscle alpha(2)-adrenoceptors whereas clonidine-induced coronary vasodilatation is mediated by activation of vascular H(2) histaminergic receptors. Accordingly, endothelial alpha(2)-adrenoceptors does not seem to play a major role in coronary flow response induced by clonidine.

Rhythmic properties of neurons in the rostral ventrolateral medulla of the rat in vitro: effects of clonidine.

The rostral ventrolateral medulla (RVLM) is thought to be the main central site for generation of tonic sympathetic activity. In the rat in vitro slice preparation, we used intracellular recordings to identify different populations of neurons in the RVLM: 43 spontaneously active neurons with regular (R) or irregular (I) patterns of spike firing and 10 silent neurons. The degree of regularity was quantified by the coefficient of variation (CV = SD/mean) of interspike interval durations, as well as by the rhythmic properties of the spike autospectrum and autocorrelation. The distribution of CVs was clustered: R and I neurons were defined as those with CVs 12% (n = 22), respectively. The R-type and I-type neurons resemble the type II and type I neurons, respectively, which were previously characterized in the RVLM in vivo as barosensitive and bulbospinal. Both types may be important in generation of sympathetic tone. Clonidine (1-100 microM) was applied to 10 R-type neurons and 16 I-type neurons. The firing of 21/26 was depressed to the point of silence. However, 18/26 neurons were excited earlier in the perfusion. The later depression of firing occurred in both I and R neurons and in different cases was associated with either hyperpolarization or depolarization.

Perineural alpha(2A)-adrenoceptor activation inhibits spinal cord neuroplasticity and tactile allodynia after nerve injury.

BACKGROUND: Nerve injury in animals increases alpha(2)-adrenoceptor expression in dorsal root ganglion cells and results in novel excitatory responses to their activation, perhaps leading to the phenomenon of sympathetically maintained pain. In contrast to this notion, peripheral alpha(2)-adrenoceptor stimulation fails to induce pain in patients with chronic pain. We hypothesized that alpha(2) adrenoceptors at the site of nerve injury play an inhibitory, not excitatory role. METHODS: Partial sciatic nerve ligation was performed on rats, resulting in a reduction in withdrawal threshold to tactile stimulation. Animals received perineural injection at the injury site of clonidine, saline, or clonidine plus an alpha(2)-adrenergic antagonist, and withdrawal threshold was monitored. Immunohistochemistry was performed on the sciatic nerve ipsi- and contralateral to injury and on the spinal cord. RESULTS: Clonidine reduced this hypersensitivity in a dose-dependent manner, and this was blocked by an alpha(2A)-preferring antagonist. Perineural clonidine injection had a slow onset (days) and prolonged duration (weeks). Systemic or intrathecal clonidine, or transient neural blockade with ropivacaine, had short lasting or no effect on hypersensitivity. alpha(2A)-adrenoceptor immunostaining was increased near the site of peripheral nerve injury, both in neurons and in immune cells (macrophages and T lymphocytes). Phosphorylated cAMP response element binding protein (pCREB) in lumbar spinal cord was increased ipsilateral to nerve injury, and this was reduced 1 week after perineural clonidine injection. CONCLUSIONS: These data suggest that peripheral alpha(2) adrenoceptors are concentrated at the site of peripheral nerve injury, and their activation receptors produce long-lasting reductions in abnormal spinal cord gene activation and mechanical hypersensitivity.

Clonidine toxicity revisited.

The incidence of clonidine overdose is increasing, yet there is a paucity of new information regarding treatment options for clonidine toxicity. Reported treatment approaches vary widely, demonstrating the lack of science on which current treatment is based. Available research needs to be reassessed. Neurotransmitters, receptors, endogenous opioids, and baseline sympathetic tone determine the clinical response to clonidine as well as the potential response to drug therapy following clonidine overdose. This article reviews aspects of clonidine toxicity that need to be further investigated. Multicenter research trials will be required to evaluate new treatment options.

Agmatine, an endogenous ligand at imidazoline binding sites, does not antagonize the clonidine-mediated blood pressure reaction.

Since agmatine has been identified as a clonidine displacing substance (CDS), the aim of this study was to investigate whether agmatine can mimic CDS-induced cardiovascular reactions in organ bath experiments, pithed spontaneously hypertensive rats (SHR) and anaesthetized SHR. Intravenously-administered agmatine significantly reduced the blood pressure and heart rate of anaesthetized SHR at doses higher than 1 and 3 mg kg(-1), respectively. These effects are probably mediated via central mechanisms, since there was an approximate 8 fold rightward shift of the dose-response curve in the pithed SHR (indicating a weakened cardiovascular effect). Moreover, in organ bath experiments, agmatine failed to alter the contractility of intact or endothelium-denuded aortal rings. When agmatine was administered i.c.v. to anaesthetized SHR, blood pressure was increased without any alteration of heart rate, whereas blood pressure was unchanged and heart rate was increased after injection into the 4th brain ventricle. This suggests that haemodynamic reaction patterns after central application are related to distinct influences on central cardiovascular mechanisms. Agmatine reduces noradrenaline release in pithed SHR while alpha(2)-adrenoceptors are irreversibly blocked with phenoxybenzamine, but not while I(1)-binding sites are selectively blocked with AGN192403. This suggests that agmatine may modulate noradrenaline release in the same way that clonidine does, i.e. via imidazoline binding sites; this involves a reduction in sympathetic tone which in turn reduces blood pressure and heart rate. Finally, CDS-like cardiovascular activity appears not to be due to agmatine, since (i) blood pressure in anaesthetized SHR is decreased by agmatine and clonidine, and (ii) agmatine did not antagonize the blood pressure reaction to clonidine in pithed or anaesthetized SHR.

Ventilatory effects of alpha2-adrenoceptor blockade in awake goats.

We sought to determine the extent to which alpha2-adrenoceptor (alpha2-AR) pathways exert a tonic influence on respiratory rhythm under normal physiological conditions. The ventilatory effects of alpha2-AR blockade with SKF-86466 were examined in awake adult goats. The antagonist effect of SKF-86466 at alpha(2)-ARs was evident both as a reversal of the ventilatory disturbances produced by the alpha2-AR agonist clonidine (5 microg/kg IV) and as a rightward shift in the potency of the agonist-evoked response after SKF-86466 pretreatment. Systemic administration of SKF-86466 in cumulative doses (25-250 microg/kg) or as a single bolus injection (500 microg/kg) produced dose-dependent increases in breathing in all animals. The excitatory effect was primarily mediated by increases in respiratory frequency that persisted for at least 120 min after injection (500 microg/kg). The time course and magnitude of the ventilatory response to SKF-86466 was similar in carotid body denervated animals. The ventilatory response to isocapnic hypoxia but not normoxic hypercapnia was significantly elevated after SKF-86466 administration (500 microg/kg). SKF-86466 (25-1000 microg/kg) produced dose-dependent increases in heart rate in all animals but did not significantly change mean arterial blood pressure at any dose. The results demonstrate that alpha2-AR pathways exert a tonic inhibitory influence on respiratory rhythm in the awake goat.

Clonidine-induced antinociception and locomotor hypoactivity are reduced by dexamethasone in mice.

The effects of dexamethasone pretreatment on clonidine-induced antinociception and locomotor hypoactivity were investigated in mice. In the hot-plate and the tail-flick tests, dexamethasone administered intraperitoneally at a dose of 1 mg kg(-1), 30 or 60 min before clonidine, reduced clonidine antinociception in both tests and reduced clonidine-induced locomotor hypoactivity in the activity cage. When administered 15 min before clonidine, dexamethasone had no effect on clonidine antinociception. A higher dexamethasone dose (10 mg kg(-1)) induced the same effects observed at a dose of 1 mg kg(-1) in the hot-plate and the tail-flick tests, but the former dose had a stronger effect on locomotor hypoactivity. Dexamethasone (10 ng/mouse) administered intracerebroventricularly 30 min before clonidine was also able to reduce both clonidine-induced antinociception and locomotor hypoactivity. The protein synthesis inhibitor, cycloheximide, administered intraperitoneally at the dose of 10 mg kg(-1), 2 h before clonidine, was able to prevent dexamethasone effects on clonidine-induced antinociception. The glucocorticoid receptor antagonist RU-38486, administered intracerebroventricularly at the dose of 1 ng/mouse, was also able to block dexamethasone effects on clonidine-induced antinociception and locomotor hypoactivity, whereas both cycloheximide and RU-38486 per se did not influence pain sensitivity or locomotor activity. These results suggest that the dexamethasone effects on clonidine-induced antinociception and locomotor hypoactivity depend on the stimulating effects that dexamethasone exert, on the protein synthesis via the glucocorticoid receptor in the brain.

Norepinephrine release from spinal synaptosomes: auto-alpha2 -adrenergic receptor modulation.

BACKGROUND: Clonidine produces analgesia after spinal injection by activating alpha2-adrenergic receptors. Recently, clonidine has been demonstrated to increase spinal release of norepinephrine (NE) in vivo, in contrast to that anticipated by classic presynaptic autoinhibition. The purpose of the current study was to determine if clonidine could inhibit release of NE in a preparation of spinal cord tissue lacking synaptic circuits. METHODS: Crude synaptosomes were prepared from male Sprague-Dawley rat spinal cord, loaded with [3H]NE, and stimulated by potassium chloride to release [3H]NE. Samples were incubated with clonidine in the absence or presence of various inhibitors. To study the effect of alpha2a-adrenergic receptor subtypes, some animals were pretreated with an oligodeoxynucleotide (ODN) composed of a sense or antisense sequence to a portion of this receptor. RESULTS: Potassium chloride produced a concentration-dependent increase in [3H]NE release, and this release was inhibited by clonidine with a concentration producing 50% maximal inhibition (IC50) of 1.3 microm. The effect of clonidine was inhibited by the alpha2-adrenergic antagonists, yohimbine and idazoxan, but not by alpha1-adrenergic, muscarinic, or opioid antagonists. Intrathecal pretreatment with antisense ODN to alpha2A-adrenergic receptors reduced alpha2A-adrenergic receptor protein expression compared with sense ODN control and also reduced clonidine-induced inhibition of [3H]NE release. CONCLUSIONS: These data demonstrate the existence of classic autoinhibitory alpha2-adrenergic receptors in the spinal cord, probably of the alpha2Asubtype. They further suggest that clonidine-induced stimulation of spinal NE release must occur from indirect actions, presumably due to activation of a spinal circuit.