Genetic alteration of the alpha2-adrenoceptor subtype c in mice affects the development of behavioral despair and stress-induced increases in plasma corticosterone levels.

alpha2-Adrenoceptors (alpha2-AR) modulate many central nervous system functions, such as regulation of sympathetic tone, vigilance, attention, and reactivity to environmental stressors. Three alpha2-AR subtypes (alpha2A, alpha2B, and alpha2C) with distinct tissue-distribution patterns are known to exist, but the functional significance of each subtype is not clear. Since specific, alpha2-AR subtype-selective pharmacological probes are not available, mice with genetically altered alpha2C-AR expression were studied in order to investigate the possible involvement of the alpha2C-AR in physiological and behavioral responses to acute and repeated stress. A modified version of Porsolt's forced swimming test was used to assess the possible effects of altered alpha2C-AR expression on the development of behavioral despair. alpha2C-Overexpression increased and the lack of alpha2C-AR (alpha2C-KO) decreased the immobility of mice in the forced swimming test, ie alpha2C-AR expression appeared to promote the development of behavioral despair. In addition, alpha2C-KO was associated with attenuated elevation of plasma corticosterone after different stressors, and overexpression of alpha2C-ARs was linked with increased corticosterone levels after repeated stress. Moreover, the brain dopamine and serotonin balance, but not norepinephrine turnover, was dependent on alpha2C-AR expression, and the expression of c-fos and junB mRNA was increased in alpha2C-KO mice. Since alpha2C-KO produced stress-protective effects, and alpha2C-AR overexpression seemed to promote the development of changes related to depression, it is suggested that a yet-to-be developed subtype-selective alpha2C-AR antagonist might have therapeutic value in the treatment of stress-related neuropsychiatric disorders.

Effects of repeated low dose administration and withdrawal of haloperidol on sexual behaviour of male rats.

Neuroleptics are known to cause anhedonia and attenuate sexual behaviour at therapeutic doses in humans. These effects are assumed to result from the dopamine antagonism of the drugs. It has been observed that a mixed dopamine D1/D2 antagonist, haloperidol, may cause a reduction in the number of intromissions required to achieve ejaculation. On the other hand, dopamine antagonists are considered unable to modify sexual behaviour once the copulatory sequence is initiated. In this study, male rats received low doses of haloperidol (30 or 60 microg/kg) before the investigation of sexual behaviour in five consecutive days and the mating test was repeated after withdrawal periods of four and five days. Haloperidol dose-dependently reduced intromission frequency, and this effect was maintained for four days after withdrawal. Ejaculation latency was reduced in all groups, including controls. The results indicate that at low doses haloperidol dose-dependently reduces intromission frequency, and the effect of a repeated dosage may persist several days after cessation of medication.

Comparison of the effects of acute and subchronic administration of atipamezole on reaction to novelty and active avoidance learning in rats.

The effects of an alpha2-adrenoceptor antagonist, atipamezole, on exploratory behaviour in a novel environment, spontaneous motor activity and active avoidance learning were studied after acute injection and continuous infusion (0.1 mg/kg h) for 24 h and 6-9 days in rats. The effects of atipamezole on biogenic amines and their main metabolites in brain were studied after an acute injection (0.3 mg/kg s.c.) and continuous infusion (0.1 mg/kg h) for 24 h and 10 days. The level of central alpha2-adrenoceptor antagonism and the drug concentration in blood and in the brain were measured after continuous infusion for 24 h and 10 days. In behavioural tests, atipamezole had no effect on spontaneous motor activity at any of the doses studied. However, after both acute administration and continuous 24-h infusion, atipamezole decreased exploratory behaviour in a staircase test, but no longer after 6 days of continuous infusion. Acute administration of atipamezole impaired performance in active avoidance learning tests causing a learned helplessness-like behaviour. When the training was started after 7 days of continuous infusion, atipamezole significantly improved active avoidance learning. There was a significant increase in the metabolite of noradrenaline (NA), 3-methoxy-4-hydroxyphenylethyleneglycol sulphate (MHPG-SO4), after 24 h but not any longer after 10 days of continuous atipamezole infusion, although the extent of central alpha2-adrenoceptor antagonism was unchanged and the atipamezole concentration present in brain was even elevated at 10 days compared to levels after 24-h infusion. In conclusion, these results reveal that acute and subchronic atipamezole treatments have different and even opposite effects on behaviour in novel, stressful situations. After acute treatment, atipamezole potentiates reaction to novelty and stress, causing a decrease in exploratory activity and impairment in shock avoidance learning. After subchronic treatment, there was no longer any effect on exploratory behaviour and, in fact, there was an improvement in the learning of a mildly stressful active avoidance test. The changes in behaviour occurred in parallel with attenuation in the MHPG-SO4-increasing effect, thus the suppressed behaviour in the present test conditions after acute atipamezole injection is associated with a major increase in central NA release. The results support the role of alpha2-adrenoceptors and noradrenergic system in reactions both to novelty and stress and have possible implications in cognitive functions as well as in depression.

D-amphetamine and L-5-hydroxytryptophan-induced behaviours in mice with genetically-altered expression of the alpha2C-adrenergic receptor subtype.

Three human and mouse genes encoding alpha2-adrenoceptor subtypes (alpha2A, alpha2B, and alpha2C) have been cloned. The alpha2C-adrenoceptor is the most abundant alpha2-adrenoceptor subtype in the striatum and modulates metabolism of both dopamine and serotonin. To investigate the possible involvement of the alpha2C-adrenoceptor subtype in behaviours regulated by dopamine and serotonin, two strains of genetically-engineered mice were examined. One had a targeted inactivation of the alpha2C-adrenoceptor gene, and the other had tissue-specific over-expression of alpha2C-adrenoceptors. The locomotor activity of the mice was evaluated after stimulation with D-amphetamine, and the behavioural serotonin syndrome and head twitches were investigated after L-5-hydroxytryptophan treatment. In addition, the effects of D-amphetamine and L-5-hydroxytryptophan were studied after pretreatment with dexmedetomidine, a subtype-nonselective alpha2-adrenoceptor agonist. The lack of alpha2C-adrenoceptor expression increased and the over-expression of alpha2C-adrenoceptors decreased the response to D-amphetamine stimulation. The effect of alpha2C-adrenoceptor gene inactivation was more prominent in D-amphetamine-treated males than in females. Dexmedetomidine inhibited D-amphetamine-induced hyperlocomotion and the L-5-hydroxytryptophan-induced serotonin syndrome, but the inhibition was attenuated in mice lacking alpha2C-adrenoceptors. However, the head twitches induced by L-5-hydroxytryptophan were effectively inhibited by dexmedetomidine in all studied mice, which suggests that alpha2A-adrenoceptors mediate the inhibition of the head twitch response. The results lend further support to the proposed existence of functionally distinct alpha2-adrenoceptor subtypes that can serve as new and specific therapeutic targets in various neuropsychiatric diseases.

Adrenergic alpha2C-receptors modulate the acoustic startle reflex, prepulse inhibition, and aggression in mice.

Studies on animal models of stress, anxiety, aggression, and sensorimotor gating have linked specific monoamine neurotransmitter abnormalities to the cognitive and behavioral disturbances associated with many affective neuropsychiatric disorders. Although alpha2-adrenoceptors (alpha2-ARs) have been suggested to have a modulatory role in these disorders, the specific roles of each alpha2-AR subtype (alpha2A, alpha2B, and alpha2C) are largely unknown. The restricted availability of relevant animal models and the lack of subtype-selective alpha2-AR drugs have precluded detailed studies in this area. Therefore, transgenic mice were used to study the possible role of the alpha2C-AR subtype in two well established behavioral paradigms: prepulse inhibition (PPI) of the startle reflex and isolation-induced aggression. The alpha2C-AR-altered mice appear grossly normal, but subtle changes have been observed in their brain dopamine (DA) and serotonin (5-HT) metabolism. In this study, the mice with targeted inactivation of the gene encoding alpha2C-ARs (alpha2C-KO) had enhanced startle responses, diminished PPI, and shortened attack latency in the isolation-aggression test, whereas tissue-specific overexpression of alpha2C-ARs (alpha2C-OE) was associated with opposite effects. Correlation analyses suggested that both the magnitude of the startle response and its relative PPI (PPI%) were modulated by the mutations. In addition, the differences in PPI, observed between drug-naive alpha2C-OE mice and their wild-type controls, were abolished by treatment with a subtype nonselective alpha2-agonist and antagonist. Thus, drugs acting via alpha2C-ARs might have therapeutic value in disorders associated with enhanced startle responses and sensorimotor gating deficits, such as schizophrenia, attention deficit disorder, post-traumatic stress disorder, and drug withdrawal.

Evaluation of the effects of a specific alpha 2-adrenoceptor antagonist, atipamezole, on alpha 1- and alpha 2-adrenoceptor subtype binding, brain neurochemistry and behaviour in comparison with yohimbine.

In the present study we evaluated the alpha 1- and alpha 2-adrenoceptor subtype binding, central alpha 2-adrenoceptor antagonist potency, as well as effects on brain neurochemistry and behavioural pharmacology of two alpha 2-adrenoceptor antagonists, atipamezole and yohimbine. Atipamezole had higher selectivity for alpha 2- vs. alpha 1-adrenoceptors than yohimbine regardless of the subtypes studied. Both compounds had comparable affinity for the alpha 2A-, alpha 2C- and alpha 2B-adrenoceptors, but yohimbine had significantly lower affinity for the alpha 2D-subtype. This may account for the fact that significantly higher doses of yohimbine than atipamezole were needed for reversal of alpha 2-agonist (medetomidine)-induced effects in rats (mydriasis) and mice (sedation and hypothermia). The effect on central monoaminergic activity was estimated by measuring the concentrations of transmitters and their main metabolites in whole brain homogenate. At equally effective alpha 2-antagonising doses in the rat mydriasis model, both drugs stimulated central noradrenaline turnover (as reflected by increase in metabolite levels) to the same extent. Atipamezole increased dopaminergic activity only slightly, whereas yohimbine elevated central dopamine but decreased central 5-hydroxytryptamine turnover rates. In behavioural tests, atipamezole (0.1-10 mg/kg) did not affect motor activity but stimulated food rewarded operant (FR-10) responding (0.03-3 mg/kg) whereas yohimbine both stimulated (1 mg/kg) and decreased (> or = 3 mg/kg) behaviour in a narrow dose range in these tests. In the staircase test, both antagonists increased neophobia, but in the two compartment test only yohimbine (> or = 3 mg/kg) decreased exploratory behaviour. The dissimilar effects of the antagonists on neurochemistry and behaviour are thought to be caused by non alpha 2-adrenoceptor properties of yohimbine. In conclusion, the alpha 2-antagonist atipamezole blocked all alpha 2-adrenoceptor subtypes at low doses, stimulated central noradrenergic activity and had only slight effects on behaviour under familiar conditions, but increased neophobia. The low affinity for the alpha 2D-adrenoceptor combined with its unspecific effects complicates the use of yohimbine as pharmacological tool to study alpha 2-adrenoceptor physiology and pharmacology.

Genetic alteration of alpha 2C-adrenoceptor expression in mice: influence on locomotor, hypothermic, and neurochemical effects of dexmedetomidine, a subtype-nonselective alpha 2-adrenoceptor agonist.

alpha 2-Adrenergic receptors (alpha 2-ARs) regulate many physiological functions and are targets for clinically important antihypertensive and anesthetic agents. Three human and mouse genes encoding alpha 2-AR subtypes (alpha 2A, alpha 2B, and alpha 2C) have been cloned. We investigated the involvement of the alpha 2C-AR in alpha 2-adrenergic pharmacology by applying molecular genetic techniques to alter the expression of alpha 2C-AR in mice. The effects of dexmedetomidine, a subtype-nonselective alpha 2-AR agonist, on monoamine turnover in brain and on locomotor activity were similar in mice with targeted inactivation of the alpha 2C-AR gene and in their controls, but the hypothermic effect of the alpha 2-AR agonist was significantly attenuated by the receptor gene inactivation. Correspondingly, another strain of transgenic mice with 3-fold overexpression of alpha 2C-AR in striatum and other brain regions expressing alpha 2C-AR showed normal reductions in brain monoamine metabolism and locomotor activity after dexmedetomidine, but their hypothermic response to the alpha 2C-AR agonists was significantly accentuated. The hypothermic effect of alpha 2-AR agonists thus seems to be mediated in part by alpha 2C-AR. Some small but statistically significant differences between the strains were also noted in brain dopamine metabolism. Lack of alpha 2C-AR expression was linked with reduced levels of homovanillic acid in brain, and mice with increased alpha 2C-AR expression had elevated concentrations of the dopamine metabolite compared with their controls.

Continuous alpha 2-adrenoceptor blockade by atipamezole decreases neocortical high-voltage spindle activity in rats.

The present study investigates the effects of a subchronic continuous infusion of atipamezole, a potent and selective alpha 2-adrenoceptor antagonist, on neocortical high-voltage spindle (HVS) activity in rats. Six days' subcutaneous infusion of atipamezole (0.125 mg/kg per h) with osmotic minipumps decreased HVS activity significantly. The HVS activity-decreasing effect of atipamezole persisted at the same level throughout the infusion. A single subthreshold dose of an alpha 2-adrenoceptor agonist, guanfacine (0.001 mg/kg i.p.), did not affect HVS activity either before or after the continuous atipamezole treatment. The central alpha 2-adrenoceptor blocking effect of atipamezole (0.1 mg/kg per h s.c.) was confirmed to be at the same level after one, three or seven days' infusion, as assessed by measuring the antagonism of detomidine-induced mydriasis in the rat. The serum concentration of atipamezole (0.1 mg/kg per h s.c.) increased slightly from day 3 (37 +/- 11 ng/ml) to day 7 (45 +/- 4 ng/ml). In conclusion, the results of the study suggest that the suppressant effects of atipamezole on neocortical high-voltage spindle activity are preserved during subchronic continuous treatment. In addition, alpha 2-adrenoceptor blockade, as measured in the rat mydriasis model, persists at the same level during subchronic infusion.

Brain regional and adrenal monoamine concentrations and behavioral responses to stress in alcohol-preferring AA and alcohol-avoiding ANA rats.

The concentrations of monoamines, precursors and metabolites in various brain regions and the levels of catecholamines in the adrenal glands were determined from naive rats of the AA and ANA lines, and from ones immediately after an escapable shock test. The brain determinations were made with a new step-gradient ion-pair elution method on a reversed phase column and coulometric detection. Several significant differences were observed in the amine concentrations, largely confirming and extending the findings made before the genetic revitalization of the lines: in particular, the AAs, unlike other alcohol-preferring rodents, had higher 5-hydroxytryptamine concentrations. The AA rats tended to have smaller changes than the ANAs in brain aminergic systems and had significantly less change in adrenal epinephrine and dopamine levels after the shock test. The AAs were consistently found to be less active than ANAs in this shock test and in a warm-water swim test, but whether this was a cause or an effect of their brain and adrenal changes could not be determined. Our behavioral results might suggest a reduced reaction of the alcohol-preferring rats to aversive stimulation.

Behavioral and color variations between rat lines developed for differential alcohol sensitivity.

The AT and ANT rat lines, outbred for differential sensitivity to ethanol-induced motor impairment, also show a difference in their sober behavior. It is not manifested in an escapable-shock test or an amphetamine-stress test, but is shown as significantly more activity by the alcohol-insensitive ATs in a low-stress (33 degrees C) modification of the forced-swimming test. The correlation between alcohol sensitivity and swimming-test activity is, however, not significant in unselected Long Evans rats. Differences in coat color have also developed in the AT and ANT lines; it was possible to estimate whether these changes are independent of ethanol sensitivity with computer simulations. They showed, for example, that the probability of the observed loss of the agouti color in the ATs being by chance is about 0.53 and the probability is nearly 0.50 for there being a spurious line difference, i.e., unrelated to ethanol sensitivity, in which only line had lost the agouti allele. More generally, these and other simulations showed that permanently maintaining selected lines is not the optimal method for finding genetically-based factors related to ethanol sensitivity or other characteristics for which lines have been developed. Nor is either revitalization or replicate lines optimal. The best method apparently would be perpetual restarting of lines.