Carbamazepine pharmacokinetics-pharmacodynamics in genetically epilepsy-prone rats

Carbamazepine produces dose-related anticonvulsant effects in epilepsy models including the genetically epilepsy-prone rat (GEPR) model and the rat maximal electroshock model. Dose-response relationships are quantitatively different among the models. Against electroshock seizures in Sprague-Dawley rats the ED50 dose is 7.5 mg/kg whereas the ED50 against audiogenic seizures in severe seizure GEPRs (GEPR-9s) is 3 mg/kg. In contrast, the ED50 in moderate seizure GEPRs (GEPR-3s) is 25 mg/kg. The present study was designed to ascribe dose-response differences among the three rat strains to pharmacokinetic or pharmacodynamic factors. After systemic carbamazepine, pharmacokinetic studies revealed differences in area under the concentration-vs.-time curve. In other experiments, carbamazepine-induced serotonin release from hippocampus was used as a pharmacodynamic marker. In a concentration-controlled design using intracerebral microdialysis, hippocampal carbamazepine infusions produced similar concentration-response relations for the three rat strains. These data support the hypothesis that dose-response differences among the three rat strains are primarily pharmacokinetic in nature.     

Alpha 1- and alpha 2-adrenoceptors in rat cerebral cortex: effects of neonatal treatment with 6-hydroxydopamine

Neonatal 6-hydroxydopamine treatment was used to destroy the noradrenergic nerve terminals in rat cerebral cortex and thus give some insight into the in vivo regulation of alpha-adrenoceptor subtypes, which in turn provides information concerning the anatomical localization of alpha 1- and alpha 2-adrenoceptors. Treatment of rats in the neonatal period with 6-OHDA causes an irreversible decrease in noradrenaline levels of the cerebral cortex compared to controls. Differences in [3H]clonidine and [3H]prazosin binding in the cerebral cortex occurred which varied depending upon the time elapsed between denervation and the binding assay. In rats aged 7-14 days there was a 20% decrease in the number of alpha 2-adrenoceptors and a slight increase in alpha 1-adrenoceptors. In older rats (45-50 day old) both types of alpha adrenoceptors were increased. Results of this study indicate that alpha 2-adrenoceptors located on presynaptic noradrenergic terminals represent only a small proportion of the total alpha 2-adrenoceptors in rat cerebral cortex. Increases in the binding capacity after 67-OHDA treatment indicate an up-regulation phenomenon affecting alpha 1-adrenoceptors and alpha 2-adrenoceptors located on structures other than noradrenergic nerve endings.     

Phenotypic expression of the systemic toxicity of cocaine in genetically epilepsy-prone rats

The purpose of the present investigation was to determine whether the sensitivity to systemic toxic effects of cocaine is altered in genetically epilepsy-prone rats (GEPRs). Moderate seizure (GEPR-3) and severe seizure (GEPR-9) rats, and the control strain, Sprague-Dawley rats, 10 weeks of age, were lightly anesthetized with halothane and nitrous oxide. Following surgical preparation and stabilization, the animals were given a constant intravenous infusion of cocaine (4 mg/kg per min) until death. Blood pressure, ECG, and EEG were monitored continuously throughout the experiment. Cocaine doses required to produce seizures (i.e., epileptiform activity on the EEG) were not significantly different between GEPRs and control rats (16.8+/-0.6 mg/kg in GEPR-3, 18.7+/-0.7 mg/kg in GEPR-9, and 14.7+/-1.3 mg/kg in Sprague-Dawley). Seizure duration, amplitude and the number of epileptiform bursts were also similar among the three strains. Additionally, there was no significant difference in cocaine doses that produced arrhythmias and cardiac asystole between GEPRs and control. The results indicate that genetically epilepsy-prone rats do not exhibit altered sensitivity to cocaine-induced seizures despite the marked susceptibility to sound-evoked seizures. Local anesthetic-induced seizures and acoustically-evoked seizures apparently have different underlying mechanisms.     

Role of adrenoceptor-linked signaling pathways in the regulation of CYP1A1 gene expression

Alpha2-adrenoceptor agents as well as stress affect the activity of several hepatic monoxygenases including those related to CYP1A enzymes. This study was therefore designed to assess the role of central and/or peripheral catecholamines and, in particular, of adrenoceptors in the regulation of B(alpha)P-induced cytochrome CYP1A1 expression. In order to discriminate the role of central from that of peripheral catecholamines in the regulation of CYP1A1 induction, the effect of central and peripheral catecholamine depletion using reserpine versus only peripheral catecholamine depletion using guanethidine was assessed. By using selected agonists and antagonists, the role of alpha and beta-adrenoceptors in the regulation of CYP1A1 induction was evaluated. The results showed that the central catecholaminergic system has a negative regulatory effect on 7-ethoxyresorufin O-deethylase (EROD) inducibility by benzo(alpha)pyrene (B(alpha)P), and that this may be mediated via alpha1-, alpha2- and beta-adrenoceptors. Specifically, stimulation of alpha2-adrenoceptors with dexmedetomidine and blockade of alpha1- or beta-adrenoceptors with prazosin or propranolol respectively, resulted in a further increase of EROD inducibility. Adrenoceptors were found to be involved in the regulation of the CYP1A1 gene at mRNA level. Both, reduced noradrenaline release in central nervous system induced with dexmedetomidine and central catecholamine depletion, as well as blockade of central alpha1-adrenoceptors induced with prazosin, all were associated with up-regulation of CYP1A1 expression. In contrast, stimulation of central beta-adrenoceptors with isoprenaline resulted in a down-regulation of CYP1A1 expression. Our observations indicate that drugs, which stimulate or block adrenoceptors and catecholamine release may lead to complications in drug therapy and modulate the toxicity or carcinogenicity of drugs that are substrates for the CYP1A1.     

Pretreatment with N-ethoxycarbonyl-2-ethoxy-1,2-dihydroquinoline accelerates the electroshock-induced decrease in density of beta-adrenoceptors but not functional downregulation

Intrapritoneal injection of N-ethoxycarbonyl-2-ethoxy-1,2-dihydroquinoline (EEDQ) to rats significantly reduced the density of alpha 1- and alpha 2-adrenoceptors in the cerebral cortex, without affecting beta-adrenoceptors. A single dose of EEDQ markedly accelerated the development of downregulation of beta-adrenoceptors induced by a short series of electroshocks. The accumulation of cAMP, induced by isoproterenol, was unchanged in rats treated with EEDQ, while the effect of noradrenaline was reduced, to the level observed after isoproterenol alone, indicating the attenuation of alpha-adrenoceptor function. The isoproterenol-induced accumulation of cAMP was not changed by EEDQ nor electroshock, not by the combined treatment. The stimulatory action of noradrenaline was reduced after EEDQ alone or in combination with electroshock, but the effect of electroshock alone was insignificant. The results suggest that the acceleration of downregulation of beta-adrenoceptors after combined treatment with an alpha-adrenoceptor blocking agent is related to elimination of the alpha-adrenergic potentiation of accumulation of cAMP mediated by beta-adrenoceptors.     

The influence of injection of oestradiol to female rats on changes in alpha 2- and beta-adrenoceptor function induced by repeated administration of desipramine or electroconvulsive shock

Repeated daily administration to female rats of either an electroconvulsive shock (110 V, 1 sec) or desipramine (DMI; 5 mg/kg x 2) caused a progressive decrease in presynaptic alpha 2-adrenoceptor function assessed by the hypoactivity (sedation) response to clonidine (0.5 mg/kg). This reduction was maximal after approximately seven electroshocks or 8-12 days of injection of DMI. Daily administration of oestradiol (100 micrograms s.c.), starting one day prior to the commencement of administration of DMI or treatment with electroshock, markedly accelerated the onset of decreased hypoactivity responses to clonidine, but did not alter the maximum reduction induced by repeated injection of DMI or administration of electroshock. Injection of oestradiol alone had no effect on the responses to clonidine. Administration of DMI for 14 days decreased the number of both alpha 2- and beta-adrenoceptors in the cortex. Cortical beta-, but not alpha 2-adrenoceptors, were also decreased after 4 days of injection of DMI. Two and ten electroshocks moderately increased and decreased cortical alpha 2-adrenoceptors, respectively. beta-Adrenoceptors were also decreased by ten electroshocks, but two were without effect. Simultaneous administration of oestradiol had little influence on the changes in the binding of alpha 2- or beta-adrenoceptors induced by repeated administration of DMI or treatment with electroshock. Oestradiol increased the numbers of cortical alpha 2- and beta-adrenoceptors 3 and 15 days after injection, respectively.(ABSTRACT TRUNCATED AT 250 WORDS)     

The role of alpha 2-adrenoceptors in the mechanism of action of antidepressant drugs

Chronic treatment of rats with imipramine or electroconvulsive shock evokes a significant decrease in binding of [3H]-clonidine to homogenates from the rat brain cerebral cortex. The decrease in binding is accompanied with a functional hyposensitivity of alpha 2-adrenoceptors as measured by attenuation of several central effects of clonidine (e.g. attenuation of clonidine-induced hypothermia, sedation, inhibition of noradrenaline formation). The influence of these antidepressants on alpha 2-adrenoceptors may be of importance in the context of data showing that prolonged treatment with these drugs attenuates the cAMP formation after noradrenergic stimulation, and that the alpha 2-adrenoceptors together with beta-adrenoceptors contribute to the formation of the second messenger. The prolonged administration of imipramine and electroshock, as well as other antidepressant treatments, e.g. administration of the monoamine oxidase inhibitor pargyline or atypical antidepressants such as trazodone, mianserin citalopram, fluoxetine, inhibit the interaction between beta- and-alpha-adrenoceptors (with the alpha 2-subtype playing a major role) and may thus contribute to the phenomenon known as the beta-adrenoceptor down-regulation. The inhibition of the interaction between alpha and beta-adrenoceptors by antidepressants may occur in multiple ways: 1. by lowering the number of beta-adrenoceptors; 2. by lowering the number of alpha 2-receptors (e.g. after imipramine or ETC); 3. by a shift of alpha 2 receptors to a higher affinity state, which represents a desensitized form of the receptor (e.g. decrease in Kd after imipramine or citalopram); 4. by a shift to an antagonist preferring state of the receptor (e.g. after treatment with fluoxetine or trazodone); 5. by a combination of the mentioned above. The functional interaction between alpha and beta adrenoceptors is presumably not dependent upon the integrity of the noradrenergic system, as it not only occurs but also is enhanced in reserpinized or 6OHDA treated rats.     

The genetically epilepsy-prone rat

1. The genetically epilepsy-prone rat (GEPR) is a valuable model for investigating mechanisms involved in epilepsy because of the controllable nature of the convulsions and their genetic origin. 2. The GEPR exhibits audiogenic seizures (AGS) and also displays higher than normal sensitivity to convulsant drugs, kindling, electroshock and hyperthermic seizures. 3. An abnormal electroencephalographic pattern and increased thresholds for auditory evoked potentials from the cochlea and brainstem are observed in the GEPR. 4. Afterdischarge-like responses and decreased sound-induced inhibition are observed in neurophysiological recordings from neurons of the inferior colliculus (IC) in the GEPR. 5. Significant deficits of norepinephrine and serotonin are observed in many regions of the GEPR brain. 6. Increases in the number of GABAergic neurons and a reduced effectiveness of iontophoretically-applied GABA are observed in the IC of this animal. 7. GABA agonists or an excitant amino acid (EAA) antagonist block AGS susceptibility when microinjected into brainstem auditory nuclei of the GEPR up to the level of IC. 8. A GABA antagonist or an EAA agonist induces susceptibility to AGS in normal rats following microinjection into IC. An increase in EAA release in IC during AGS in the GEPR is also observed. 9. This increased release of EAA and the reduced effectiveness of GABA in IC may be important seizure initiation mechanisms in the GEPR. 10. The AGS pathway in the GEPR appears to involve the auditory nuclei up to the IC as well as the brainstem reticular formation and substantia nigra but not the entopenduncular nucleus or hippocampus.     

Effects of chronic exposure to ethanol alone and in combination with desipramine on beta-adrenoceptors of rat brain

The effect of chronic ethanol exposure alone or in combination with desipramine on agonist and antagonist binding to beta-adrenoceptors was studied in membrane preparations from rat frontal cortex and hippocampus. Ten day exposure of animals to ethanol vapor (25 mg/l) in inhalation chambers had no effect on binding properties of antagonist iodocyanopindolol (ICYP) in either brain region. However, ethanol in combination with chronic desipramine treatment prevented the reduction of beta-adrenoceptor density in frontal cortex produced by desipramine administration. Similar to its effects on antagonist binding, chronic ethanol exposure did not change the agonist isoproterenol binding characteristics measured in membranes from either rat frontal cortex or hippocampus. However, the combination of ethanol plus desipramine reduced the dissociation constant of the low affinity state of the receptor (KL) in frontal cortex from 23.1 +/- 3.7 microM in controls to 11.2 +/- 1.7 microM. Moreover, ethanol plus desipramine produced a greater decrease in the percentage of cortical receptors in the high affinity state for agonist (%RH) than did desipramine alone. This suggests that ethanol enhances desipramine-induced desensitization of beta-adrenoceptors in frontal cortex in spite of the prevention of reduction in density of the receptors. In hippocampal membranes, ethanol together with desipramine prevented desipramine-induced changes in agonist binding characteristics, i.e. the decrease in KH (dissociation constant from high affinity state of the receptor) and the consequent enhancement in KL/KH ratio. Thus, chronic exposure to relatively low concentrations of ethanol partially prevents effects of desipramine on beta-adrenoceptors.     

Receptor subtype abundance as a tool for effective intracellular signalling

The regulation of heart function is one of the essentials for the survival of organism. Therefore, the effective mechanisms of regulation can minimize the energy requirements and improve the ability to react to different needs on time and appropriately. Two receptor types, beta-adrenoceptors and muscarinic receptors, with almost antagonistic function, are "basic regulators" of the heart parameters. It is relevant to mention that beside the main adrenoceptors and muscarinic receptors subtypes (beta(1)- and M(2)-subtype), other minor subtypes that regulate heart function, i.e. beta-, beta-adrenoceptors, alpha(1)-adrenoceptors and minor subtypes of muscarinic receptors (M(1), M(3) and M(5)) are present in the heart. In this regard is intriguing that just two catecholamines (adrenaline, noradrenaline) have many "targets"--receptors that differ so much in the functional consequences of their activation: while beta(1)- and beta(2)-adrenoceptors cause cardiostimulation, beta(3)-adrenoceptors are responsible for cardioinhibition and alpha(1)-adrenoceptors contribute to enhanced inotropy. Similarly, some data show that other muscarinic receptors than M(2) muscarinic subtype, are expressed in the heart and these minor subtype(s) can contribute to the heart regulation in similar way as beta(3)-adrenoceptors to the catecholamine action. Taken together, regulation of heart function through different receptor subtypes and using homologous and heterologous regulation can represent an effective tool for coping with permanently changing environmental conditions.     

Ten day administration of desipramine produces an increase in KL/KH for beta-receptors in rat hippocampus

The action of chronic desipramine administration on agonist and antagonist binding to beta-adrenoceptors was investigated in rat frontal cortex and hippocampus. Ten day treatment with desipramine (10 mg/kg per day i.p.) resulted in a significant 34% decrease in the density of beta-adrenoceptors in membranes from rat frontal cortex but not hippocampus. The KD values for antagonist (iodocyanopindolol) did not change in either tissue after the desipramine treatment. Desipramine had no effect on the affinity of the agonist isoproterenol in frontal cortex. The percentage of receptors in the high affinity state for isoproterenol (% RH) decreased by 11% in the membranes from frontal cortex after desipramine administration. In contrast, in hippocampal membranes, desipramine treatment produced a decrease in KH (dissociation constant for isoproterenol binding to the high affinity state) and consequently a 137% increase in the ratio of dissociation constants for isoproterenol to the low and high affinity conformations (KL/KH; an putative index of coupling). This observation is new and suggests possible supercoupling of beta-adrenoceptors in hippocampus during desipramine treatment. Thus, in addition to desensitization of beta-adrenoceptors in rat frontal cortex ten day administration of desipramine causes a change compatible with supercoupling of beta-receptors in hippocampal membranes.     

The effects of electroconvulsive shock or imipramine on subtypes of alpha 1-adrenoceptors in the frontal cortex of the rat.

The effects of repeated treatment (14 days) with electroconvulsive shock (ECS) or imipramine on binding sites on alpha 1-adrenoceptors in the rat were studied. The binding of [3H]prazosin studied with WB4101 and phentolamine, as binding inhibitors, showed the existence of two subtypes of alpha 1-adrenoceptor (alpha 1A and alpha 1B). Proportions of the alpha 1A and alpha 1B binding sites were about 3:7 in the frontal cortex and 9:1 in the hippocampus. Pretreatment of the membranes with chlorethylclonidine (CEC) almost abolished the alpha 1B binding sites. Inhibition of the binding of [3H]prazosin studied with antidepressants (imipramine, desipramine, maprotiline and mianserin) showed that these drugs bound to alpha 1-adrenoceptors with low affinity, in an apparent monophasic manner. The characteristics of the alpha 1A and alpha 1B binding sites were studied by the binding assay with [3H]prazosin, in the presence of a small concentration (2 nM) of WB4101 to mask the alpha 1A binding sites, as well as the assay without WB4101, for the total alpha 1-adrenoceptor (alpha 1A and alpha 1B) binding. Repeated treatment with electroconvulsive shock increased but that with imipramine decreased, the density of the alpha 1B binding sites in the frontal cortex, without change of the affinity. Neither treatment affected the alpha 1A binding sites in the frontal cortex. The alpha 1-adrenoceptors (alpha 1A and alpha 1B) in the hippocampus were not affected at all by these repeated treatments. The electroconvulsive shock-induced increase in the alpha 1B binding sites in the frontal cortex of the rat could contribute to differences in clinical effects between electroconvulsive shock and antidepressant drugs.     

Alpha-adrenoceptors and monoamine contents in the cerebral cortex of the rodent Jaculus orientalis: effects of acute cold exposure

The tritiated adrenergic antagonists prazosin ([3H]PRZ) and idazoxan ([3H]IDA, or RX-781094) bind specifically and with high affinity to alpha 1- and alpha 2-adrenoceptors respectively, and were used to measure adrenoceptors in membrane preparations obtained from the cerebral cortex of Jaculus orientalis. Membrane preparations were also obtained from a group of cold exposed animals, to determine whether these adrenoceptors could be modified by a thermic stress. The density of receptors (Bmax; maximum binding capacity) and the dissociation constant (Kd 25 degrees C) were estimated by iterative modelling, and by using the procedure of Hill. After acute cold exposure (16 hr, 5 degrees C) there was a decrease in the affinity of the alpha 1-adrenoceptors, as judged by the Kd 25 degrees C for [3H]PRZ, with no changes in the Bmax. The alpha 2-sites did not show any significant changes, as revealed by [3H]IDA binding. Pretreatment of the membrane preparations from control animals with the disulfide and sulfhydryl reactives DL-dithiothreitol, 5,5'-dithiobis-(2-nitrobenzoic acid) and N-ethylmaleimide decreased specific [3H]PRZ and [3H]IDA binding, with minor changes in non-specific counts, indicating that the fixation of these ligands was to the receptor proteins. The endogenous cortical monoamine contents were also determined in the frontal cerebral cortex of these same animals, using high performance liquid chromatography with electrochemical detection. The catecholamine levels and their major metabolites were found to be stable in the cortex after the acute thermic stress, but there was a marked reduction in serotonin with a normal content in 5-hydroxyindole-3-acetic acid.     

Acute and chronic effects of methylphenidate on cortical adrenoceptors in the rat

The effects of acute and chronic treatments with methylphenidate were examined on cortical adrenoceptors. A single dose of methylphenidate increased the affinity of [3H]prazosin and [3H]idazoxan binding sites. Acute and chronic methylphenidate treatments caused a down-regulation of alpha 1-adrenoceptors. There were no changes in cortical beta-adrenoceptors measured with [3H]dihydroalprenolol nor in endogenous monoamine levels. The alterations in alpha-adrenoceptors can be attributed to an indirect action of methylphenidate possibly through an accrued release of noradrenaline or tyramine.     

Inhibitory effects of 6-hydroxydopamine on the clonic convulsions induced by electroshock and decapitation.

Effects of intraventricular injection of 6-hydroxydopamine (6-OHDA) on maximal electroshock seizures and decapitation convulsions were investigated in mice. The incidence of the clonic phase of maximal electroshock seizure was suppressed by 6-OHDA. The time course of the suppression was in good agreement with the decreased level of whole brain norepinephrine. The incidence of clonic convulsions following decapitation was also suppressed by 6-OHDA. On the contrary, the tonic phase of maximal electroshock seizure was potentiated by the treatment. The incidence of the clonic phase of maximal electroshock seizure and of decapitation convulsions were well correlated in 6-OHDA treated mice which had been exposed to maximal electroshock seizure and then decapitated. It is suggested that the central catecholaminergic mechanism concerned with the clonic phase in maximal electroshock seizure is analogous to that concerned with decapitation convulsions and different from that underlying the tonic phase of maximal electroshock seizure.     

Modulation of adrenergic receptors during left ventricular hypertrophy development and after regression by captopril.

The objective of this study was to analyze adrenergic receptors during cardiac hypertrophy development, after establishment of cardiac hypertrophy and after regression of cardiac hypertrophy by an angiotensin-converting enzyme inhibitor. Left ventricular hypertrophy (LVH) was induced by abdominal aortic stenosis. After surgery, plasma norepinephrine concentrations (PNE) and left ventricular adrenergic receptors from rat hearts subjected to aortic stenosis were assessed during cardiac hypertrophy development (at 3, 7, 15, and 30 days of aortic stenosis), once cardiac hypertrophy had been established (7 and 14 weeks after the stenosis) and after regression of cardiac hypertrophy by an antihypertensive dose (200 mg/kg/day) of captopril. The presence of LVH was observed from day 7 after stenosis. PNE had significantly increased after 15 days but returned to control values 30 days after surgery. The density of alpha1-adrenoceptors was found to decrease with development of hypertrophy. Once hypertrophy had been established, 7 weeks from stenosis, PNE was not different from control; however, the density of alpha1-adrenoceptors continued to diminish, whereas PNE and the density of beta-adrenoceptors were no different from control values. Fourteen weeks after stenosis, a significant decrease in PNE was recorded, and no change in alpha1- but an increase in beta-adrenoceptors was observed. LVH was reversed by treatment with captopril; PNE was similar in control and stenosed treated animals. The density of alpha1-adrenoceptors was decreased when compared with control animals, and no change in the density of beta-adrenoceptors was observed with treatment. In conclusion, a decrease of alpha1-adrenoceptors was associated with LVH development and earlier stages of established cardiac hypertrophy. Later stages of established cardiac hypertrophy were characterized by no change in alpha1- and an increase in beta-adrenoceptors. Treatment with captopril induced LVH regression and decreased the number of alpha1-adrenoceptors without any change in beta-adrenoceptors.     

Gene expression profiles in response to the activation of adrenoceptors in A7r5 aortic smooth muscle cells

1. Vascular adrenoceptors play an important role in vascular physiology and pathophysiology, such as hypertension, atherosclerosis and restenosis after angioplasty. To define the changes in the ene expression in vascular smooth muscle cells in response to the activation of alpha1- or beta-adrenoceptors, a DNA microarray was used. 2. First, the existence of alpha1- and beta-adrenoceptors in A7r5 aortic smooth muscle cells was confirmed by radioligand binding. Then, the inhibitory effects of phenylephrine (an alpha1-adrenoceptor agonist) and isoproterenol (a beta-adrenoceptor agonist) on the proliferation of A7r5 cells were determined by [3H]-thymidine incorporation. 3. The A7r5 cells were treated with 10 micromol/L phenylephrine or 1 micromol/L isoproterenol for 24 h and changes in gene expression were detected with the DNA microarray. Only 14 and 20 genes were identified after treatment of cells with phenylephrine and isoproterenol, respectively, and most genes displayed decreased expression. The changed genes could be grouped into five major functional categories: cell signalling/communication, cell structure/motility, cell/organism defence, gene/protein expression and metabolism. The gene expression profile in response to the activation of alpha1-adrenoceptors was very different from that following activation of beta-adrenoceptors. Interestingly, many phenylephrine-responsive genes were associated with metabolism, whereas many isoproterenol-responsive genes encoded cell signalling and structure proteins. This means that adrenoceptors may modulate multiple aspects of biological function in vascular smooth muscle cells. 4. Collectively, the activation of both alpha1-adrenoceptors (with phenylephrine) and beta-adrenoceptors (with isoproterenol) inhibited the proliferation of A7r5 cells, but microarray data revealed that the mechanisms may be different: the activation of alpha1-adrenoceptors could induce the expression of metabolic genes, resulting in the inhibition of proliferation, whereas activation of beta-adrenoceptors altered the expression of genes that encoded cell signalling and structure proteins to inhibit cell proliferation.     

The contraction mechanisms for norepinephrine in single cells prepared from tracheal smooth muscles of guinea pig of different ages.

Single cells were prepared from guinea pig tracheal smooth muscle and used in an experiment on the contraction mechanisms for norepinephrine and a study on the change of alpha 2-adrenoceptors with age. Specific bindings of [3H]p-aminoclonidine to the single cells from the tracheal smooth muscles of 6- and 40-week-old guinea pigs were saturable and analyzed by Scatchard plot. The maximum number of [3H]p-aminoclonidine binding sites was larger in the preparation from 40-week-old guinea pigs than that from 6-week-old animals, while its dissociation constant did not change with age. The amount of prostaglandin F2 alpha released from the single cells was increased by norepinephrine, not affected by phenylephrine, and reduced by an alpha 2-antagonist such as yohimbine. The amount released by norepinephrine was significantly larger in the preparation from 6-week-old guinea pigs than that from 40-week-old animals. These results suggest that the age-related decrease in the potency of norepinephrine is due to reduction in the amount of excitable prostaglandin F2 alpha released by the drug, but not to a change in the total amount of alpha 2-adrenoceptors or the dissociation constant of the drug with respect to these adrenoceptors. Furthermore, alpha 2-adrenoceptors in the tracheal smooth muscle cell play an important role in the release of prostaglandin F2 alpha and the production of contractile responses of these muscles.     

Effects of chemical sympathectomy with 6-hydroxydopamine on alpha- and beta-adrenoceptors and muscarinic cholinoceptors in rat kidney

The autonomic receptors in the rat kidney were characterized using the radioligands [3H]prazosin, [3H]clonidine, [3H]dihydroalprenolol (DHA) and [3H]quinuclidinyl benzilate (QNB). The specific binding of [3H]prazosin, [3H]clonidine, [3H]DHA and [3H]QNB to rat kidney membranes was saturable and of high affinity, and showed a pharmacological specificity as well as stereospecificity which characterized renal alpha 1-, alpha 2- and beta-adrenoceptors and muscarinic cholinoceptors, respectively. There was a relatively greater density of alpha-adrenoceptors than beta-adrenoceptors or muscarinic cholinoceptors in the rat kidney. Chemical sympathectomy of rats with 6-hydroxydopamine X HBr (6-OHDA, 50 X 2 mg/kg i.v., 24 h interval) caused a significant increase (21-56%) in the Bmax values for renal [3H]prazosin, [3H]clonidine and [3H]DHA binding at 1 and 2 weeks following the treatment, without a change in the Kd values. 6-OHDA treatment had no significant effect on the Kd and Bmax values for [3H]QNB binding at 1-3 weeks after the treatment. The norepinephrine (NE) concentration was reduced (68-76%) in the 6-OHDA-treated rat kidney. In conclusion, the present study provides biochemical evidence for the possible localization of postsynaptic alpha 1-, alpha 2- and beta-adrenoceptors and muscarinic cholinoceptors in the rat kidney and also for the regulation of these adrenoceptors by the sympathetic nervous system.     

Alpha 2-adrenoceptors as a target for formamidine pesticides: in vitro and in vivo studies in mice

While the toxicity in insects of formamidines such as chlordimeform (CDM), its demethylated metabolite DCDM, and amitraz (AMZ) appears to involve activation of an octopamine-sensitive adenylate cyclase, their mechanism of action in mammals remains elusive. There is increasing evidence, however, that alpha 2-adrenoceptors might mediate certain effects of CDM, DCDM, and AMZ. In the present study, we investigated whether formamidines can interact directly with adrenoceptors in mouse forebrain both in vitro and after in vivo administration. Formamidines were potent inhibitors of the binding of [3H]clonidine to alpha 2-adrenoceptors with IC50's of 13 microM, 29 nM, and 130 nM for CDM, DCDM, and AMZ, respectively. Binding of [3H]yohimbine was inhibited with similar potencies. All compounds also inhibited with equal (CDM) or lower potency the binding of [3H]spiperone to dopamine D2 receptors and were weak inhibitors or inactive toward alpha 1- and beta-adrenoceptors, cholinergic muscarinic, GABAA, opiate mu, benzodiazepine, and histamine 1 receptors. Administration of formamidines to mice caused a dose-dependent decrease of [3H]clonidine binding. [3H]Clonidine binding returned to control values within 5 hr following administration of CDM and DCDM, but was still significantly decreased up to 48 hr after AMZ. Among different brain regions, [3H]clonidine binding was decreased to a larger extent in cerebral cortex, hippocampus, and midbrain. In vitro and ex vivo kinetic binding studies indicated that the effect of formamidines on alpha 2-adrenoceptors was due to a decrease in affinity and not to an alteration of the density of [3H]clonidine binding sites. The results of these biochemical studies support the hypothesis that alpha 2-adrenoceptors represent an important target for formamidine neurotoxicity in mammals.