Anabolic-androgenic steroid treatment induces behavioral disinhibition and downregulation of serotonin receptor messenger RNA in the prefrontal cortex and amygdala of male mice

Nandrolone is an anabolic-androgenic steroid (AAS) that is highly abused by individuals seeking enhanced physical strength or body appearance. Supraphysiological doses of this synthetic testosterone derivative have been associated with many physical and psychiatric adverse effects, particularly episodes of impulsiveness and overt aggressive behavior. As the neural mechanisms underlying AAS-induced behavioral disinhibition are unknown, we investigated the status of serotonergic system-related transcripts in several brain areas of mice receiving prolonged nandrolone administration. Male C57BL/6J mice received 15 mg/kg of nandrolone decanoate subcutaneously once daily for 28 days, and different sets of animals were used to investigate motor-related and emotion-related behaviors or 5-HT-related messenger RNA (mRNA) levels by real-time quantitative polymerase chain reaction. AAS-injected mice had increased body weight, were more active and displayed anxious-like behaviors in novel environments. They exhibited reduced immobility in the forced swim test, a higher probability of being aggressive and more readily attacked opponents. AAS treatment substantially reduced mRNA levels of most investigated postsynaptic 5-HT receptors in the amygdala and prefrontal cortex. Interestingly, the 5-HT(1B) mRNA level was further reduced in the hippocampus and hypothalamus. There was no alteration of 5-HT system transcript levels in the midbrain. In conclusion, high doses of AAS nandrolone in male mice recapitulate the behavioral disinhibition observed in abusers. Furthermore, these high doses downregulate 5-HT receptor mRNA levels in the amygdala and prefrontal cortex. Our combined findings suggest these areas as critical sites for AAS-induced effects and a possible role for the 5-HT(1B) receptor in the observed behavioral disinhibition.

Functional and Structural Connectivity Between the Perigenual Anterior Cingulate and Amygdala in Bipolar Disorder

Objective: Abnormalities in the morphology and function of two gray matter structures central to emotional processing, the perigenual anterior cingulate cortex (pACC) and amygdala, have consistently been reported in bipolar disorder (BD). Evidence implicates abnormalities in their connectivity in BD. This study investigates the potential disruptions in pACC-amygdala functional connectivity and associated abnormalities in white matter that provides structural connections between the two brain regions in BD. Methods: Thirty-three individuals with BD and 31 healthy comparison subjects (HC) participated in a scanning session during which functional magnetic resonance imaging (fMRI) during processing of face stimuli and diffusion tensor imaging (DTI) were performed. The strength of pACC-amygdala functional connections was compared between BD and HC groups, and associations between these functional connectivity measures from the fMRI scans and regional fractional anisotropy (FA) from the DTI scans were assessed. Results: Functional connectivity was decreased between the pACC and amygdala in the BD group compared with HC group, during the processing of fearful and happy faces (p < .005). Moreover, a significant positive association between pACC-amygdala functional coupling and FA in ventrofrontal white matter, including the region of the uncinate fasciculus, was identified (p < .005). Conclusion: This study provides evidence for abnormalities in pACC-amygdala functional connectivity during emotional processing in BD. The significant association between pACC-amygdala functional connectivity and the structural integrity of white matter that contains pACC-amygdala connections suggest that disruptions in white matter connectivity may contribute to disturbances in the coordinated responses of the pACC and amygdala during emotional processing in BD.

Anxiety-induced antinociception in mice: effects of systemic and intra-amygdala administration of 8-OH-DPAT and midazolam

Rationale: Mice exhibit antinociception after a single experience in the elevated plus maze (EPM), an animal model of anxiety. Objective: This study investigated the mechanisms involved in this form of anxiety-induced antinociception. Methods: Nociception was evaluated by means of the writhing test in mice confined either to the open or enclosed arms of the EPM. The effects of systemic (naloxone, midazolam and 8-OH-DPAT) or intra-amygdala (8-OH-DPAT. NAN-190 and midazolam) drug infusions were investigated in mice previously treated i.p. with 0.6% acetic acid, an algic stimulus that induces abdominal contortions. The effects of these drugs on conventional measures of anxiety (% entries and % time in open arms) in a standard EPM test were also independently investigated. Results: Open-arm confinement resulted in a high-magnitude antinociception (minimum 85%, maximum 450%) compared with enclosed arm confinement. The opiate antagonist naloxone (1 mg/kg and 10 mg/kg) neither blocked this open arm-induced antinociception (OAIA) nor modified indices of anxiety in EPM. Administration of midazolam (0.5-2 mg/kg, s.c.) increased OAIA and produced antinociception in enclosed confined animals, as well as attenuating anxiety in the EPM. The 5-HT(1A) receptor agonist 8-OH-DPAT (0.05-1 mg/kg, s.c.) had biphasic effects on OAIA, antagonising the response at the lowest dose and intensifying it at the highest dose. In addition, low doses of this agent reduced anxiety in the EPM. Although bilateral injections of 8-OH-DPAT (5.6 nmol/0.4 mu l) or NAN-190 (5.6 nmol and 10 nmol/0.4 mu l) into the amygdala did not alter OAIA, increased anxiety was observed in the EPM. In contrast, intra-amygdala administration of midazolam (10 nmol and 30 nmol/0.4 mu l) blocked both OAIA and anxiety. Conclusions: These results with systemic and intracerebral drug infusion suggest that 5-HT(1A) receptors localised in the amygdala are not involved in the pain inhibitory processes that are recruited during aversive situations. However, activation of these receptors does phasically increase anxiety. Although the intrinsic antinociceptive properties of systemically administered midazolam confounded interpretation of its effects on OAIA, intra-amygdala injections of this compound suggest that benzodiazepine receptors in this brain region modulate both the antinociceptive and behavioural (anxiety) responses to the EPM.

Anxiogenic-like effects of mCPP microinfusions into the amygdala (but not dorsal or ventral hippocampus) in mice exposed to elevated plus-maze

Serotonin (5-HT) can either increase or decrease anxiety-like behaviour in animals, actions that depend upon neuroanatomical site of action and 5-HT receptor subtype. Although systemic studies with 5-HT(2) receptor agonists and antagonists suggest a facilitatory role for this receptor subtype in anxiety, somewhat inconsistent results have been obtained when such compounds have been directly applied to limbic targets such as the hippocampus and amygdala. The present study investigated the effects of the 5-HT(2B/2C) receptor agonist mCPP bilaterally microinjected into the dorsal hippocampus (DH: 0, 0.3 1.0 or 3.0 nmol/0.2 mu l), the ventral hippocampus (VH: 0, 0.3, 1.0 or 3.0 nmol/0.2 mu l) or the amygdaloid complex (0, 0.15, 0.5, 1.0 or 3.0 nmol/0.1 mu l) in mice exposed to the elevated plus-maze (EPM). Test sessions were videotaped and subsequently scored for conventional indices of anxiety (percentage of open arm entries and percentage of open arm time) and locomotor activity (closed arm entries). Results showed that mCPP microinfusions into the DH or VH failed to affect any behavioural measure in the EPM. However, when injected into the amygdaloid complex, the dose of 1.0 nmol of this 5HT(2B/2C) receptor agonist increased behavioural indices of anxiety without significantly altering general activity levels. This anxiogenic-like effect of mCPP was selectively and completely blocked by local injection of a behaviourally-inactive dose of SDZ SER-082 (10 nmol/0.1 mu l), a preferential 5-HT(2C) receptor antagonist. These data suggest that 5HT(2C) receptors located within the amygdaloid complex (but not the dorsal or ventral hippocampus) play a facilitatory role in plus-maze anxiety in mice. (c) 2007 Elsevier B.V. All rights reserved.

Blockade of fear-induced antinociception with intra-amygdala infusion of midazolam: Influence of prior test experience

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Alterações na evocação de uma memória aversiva ao liongo do ciclo estral de ratas: influências da transmissão gabaérgica na amígdala

GABAergic neurotransmission has been implicated in many aspects of learning and memory, as well as mood and anxiety disorders. The amygdala has been one of the major focuses in this area, given its essential role in modulating emotionally relevant memories. However, studies with male subjects are still predominant in the field. Here we investigated the consequences for an aversive memory of enhancing or decreasing GABAergic transmission in the basolateral nucleus of the amygdala (BLA). Wistar female rats were trained in the plus-maze discriminative avoidance task, in which they had to learn to avoid one of the enclosed arms where an aversive stimulus consisting of a bright light and a loud noise was given (day 1). Fifteen minutes before the test session (day 2) animals received 0,2 μL infusions of either saline solution, the GABAergic agonist muscimol (0,05 mg/ml), or the GABAergic antagonist bicuculine (0,025 mg/ml) bilaterally intra-BLA. On the test day, females in proestrous or estrous presented adequate retrieval and did not extinguish the task, while females in metestrous or diestrous presented impaired retrieval. In the first group, muscimol infusion impaired retrieval and bicuculline had no effect, suggesting naturally low levels of GABAergic transmission in the BLA of proestrous and estrous females. In the second group, muscimol infusion had no effect and bicuculline reversed retrieval impairment, suggesting naturally high levels of GABAergic transmission in the BLA of metestrous and diestous females. Additionally, proestrous and estrous females presented higher anxiety levels compared to metestrous and diestrous females, which could explain better performance of this group. On the other hand, BLA GABAergic system did not interfere with the innate fear response because drug infusions had no effect in anxiety. Thus, retrieval alterations caused by the GABAergic drugs were probably related specifically to memory processes

Damage to the central amygdala produces differential encephalic c-fos expression in the water deprivation-partial rehydration protocol

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Lesions in the central amygdala impair sodium intake induced by the blockade of the lateral parabrachial nucleus

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Lateral parabrachial nucleus and central amygdala in the control of sodium intake

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

EFFECT OF ELECTROLYTIC AND CHEMICAL LESION BY IBOTENIC ACID OF THE AMYGDALA ON SALT INTAKE

Sodium chloride intake was studied in male Holtzman rats weighing 250-300 g submitted to electrolytic and chemical lesion of the cell bodies, not fibers of the amygdaloid complex. Sodium chloride (1.5%) intake increased in animals with electrolytic lesion of the corticomedial nucleus of the amygdala. Sodium chloride (1.5%) intake increased after ibotenic acid injection into the corticomedial nucleus of the amygdala to a larger extent (26.6 +/- 9.2 to 147.6 +/- 34.6 ml/5 days). The results indicate that sodium intake response can be induced by lesions, which involved only cell bodies. The fibers of passage of the corticomedial nucleus of the amygdala produce a water intake less consistent than that induced by ibotenic acid, which is more acute. The results show that cell bodies of this region of the amygdala are involved in the control of sodium chloride intake.

Similar anxiolytic-like effects following intra-amygdala infusions of benzodiazepine receptor agonist and antagonist: Evidence for the release of an endogenous benzodiazepine inverse agonist in mice exposed to elevated plus-maze test

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)

Effect of adrenergic stimulation of the amygdaloid complex on water intake

The effect of noradrenaline, isoproterenol, phentolamine and propranolol, injected into the basolateral nuclei of the amygdala on water intake, was investigated in male Holtzman rats. The injection of noradrenaline (40 nmol) into the amygdaloid complex (AC) of satiated rats produced no change in water intake (0.05 ± 0.03 ml/1 hour). The injection of isoproterenol (40 nmol) produced an increase in water intake in sedated rats (1.93 ± 0.23 ml/1 hour). Noradrenaline injected into the AC produced a decrease in water intake in deprived rats (0.40 ± 0.19 ml/1 hour). The injection of isoproterenol into the AC of deprived rats produced no change in water intake in comparison with control (11.65 ± 1.02 and 10.92 ± 0.88 ml/1 hour, respectively). When compared with control values, phentolamine injected prior to noradrenaline blocked the inhibitory effect of noradrenaline on water intake in deprived rats (10.40 ± 1.31 ml/1 hour). Propranolol blocked the effect of isoproterenol in satiated rats (0.85 ± 0.49 ml/1 hour) and also blocked the water intake induced by deprivation (0.53 ± 0.38 ml/1 hour). In satiated and deprived animals the injection of phentolamine before hexamethonium blocked the inhibitory effect of hexamethonium on water intake. In satiated animals, when hexamethonium was injected alone, water intake was 0.39 ± 0.25 ml/1 hour and when hexamethonium was injected with phentolamine, water intake was 1.04 ± 0.3 ml/1 hour. In deprived animals, hexamethonium alone blocked water intake (0.40 ± 0.17 ml/1 hour) and when injected with phentolamine it elicited an intake of 9.7 ± 1.8 ml/1 hour. these results clearly demonstrate the participation of catecholaminergic receptors of the AC in the regulation of water intake.

Importance of the central nucleus of the amygdala on sodium intake caused by deactivation of lateral parabrachial nucleus

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

The control of sodium chloride intake: Functional relationship between hypothalamic inhibitory areas and amygdaloid complex stimulating areas

Sodium chloride intake was studied in rats submitted to different neurosurgical procedures. Intake decreased in animals submitted to bilateral destruction of the basolateral amygdaloid complex, and increased after the same animals were submitted to destruction of the anterior lateral hypothalamus, a procedure which is known to cause increased intake in intact rats. In the reverse experiment, where the anterior lateral hypothalamus was destroyed before the basolateral amygdaloid complex, the effect of increased sodium chloride intake induced by destruction of the hypothalamus overcame the decreased expected upon destruction of the amygdaloid complex. These results permit us to conclude that the hypothalamic areas which inhibit sodium chloride intake predominate over the stimulating areas of the amygdaloid complex in the control of sodium chloride intake. © 1981 ANKHO International Inc.

Consequences of pilocarpine-induced status epilepticus in immunodeficient mice

Systemic injection of pilocarpine in rodents induces status epilepticus (SE) and reproduces the main characteristics of temporal lobe epilepsy (TLE). Different mechanisms are activated by SE contributing to cell death and immune system activation. We used BALB/c nude mice, a mutant that is severely immunocompromised, to characterize seizure pattern, neurochemical changes, cell death and c-Fos activation secondarily to pilocarpine-induced SE. The behavioral seizures were less severe in BALB/c nude than in BALB/c wild type mice. However, nude mice presented more tonic clonic episodes and higher mortality rate during SE. The c-Fos expression was most prominent in the caudate-putamen, CA3 (p < 0.05), dentate gyrus, entorhinal cortex (p < 0.001), basolateral nucleus of amygdala (p < 0.01) and piriform cortex (p < 0.05) of BALB/c nude mice than of BALB/c. Besides, nude mice subjected to SE presented high number of Fluorojade-B (FJB) stained cells in the piriform cortex, amygdala (p < 0.05) and hilus (p < 0.05) in comparison with BALB/c mice. A significant increase in the level of glutamate and GABA was found in the hippocampus and cortex of BALB/c mice presenting SE in comparison to controls. However, the level of glutamate was higher in the brains of BALB nude mice than in the brains of BALB/c wild type mice, while the levels of GABA were unchanged. These results indicate that the brains of immunodeficient nude mice are more vulnerable to the deleterious effects of pilocarpine-induced SE as they present intense activation, increased glutamate levels and more cell death. Published by Elsevier B.V.