Gabaergic mechanisms of hypothalamic nuclei in the expression of conditioned fear

The amygdala, the dorsal periaqueductal gray (dPAG), and the media] hypothalamus have long been recognized to be a neural system responsible for the generation and elaboration of unconditioned fear in the brain. It is also well known that this neural substrate is under a tonic inhibitory control exerted by GABA mechanisms. However, whereas there is a growing body of evidence to suggest that the amygdala and dPAG are also able to integrate conditioned fear, it is still unclear, however, how the distinct hypothalamic nuclei participate in fear conditioning. In this work we aimed to examine the extent to which the gabaergic mechanisms of this brain region are involved in conditioned fear using the fear-potentiated startle (FPS). Muscimol, a GABA-A receptor agonist, and semicarbazide, an inhibitor of the GABA synthesizing enzyme glutamic acid decarboxylase (GAD), were used as an enhancer and inhibitor of the GABA mechanisms, respectively. Muscimol and semicarbazide were injected into the anterior hypothalamus (AHN). the dorsomedial part of the ventromedial nucleus (VMHDM), the dorsomedial (DMH) or the dorsal premammillary (PMD) nuclei of male Wistar rats before test sessions of the fear conditioning paradigm. The injections into the DMH and PMD did not produce any significant effects on FPS. On the other hand, muscimol injections into the AHN and VMHDM caused significant reduction in FPS. These results indicate that injections of muscimol and semicarbazide into the DMH and PMD fail to change the FPS, whereas the enhancement of the GABA transmission in the AHN and VMHDM produces a reduction of the conditioned fear responses. On the other hand, the inhibition of this transmission led to an increase of this conditioned response in the AHN. Thus, whereas DMH and PMD are known to be part of the caudal-most region of the medial hypothalamic defensive system, which integrates unconditioned fear, systems mediating conditioned fear select the AHN and VMHDM nuclei that belong to the rostral-most portion of the hypothalamic defense area. Thus, distinct subsets of neurons in the hypothalamus could mediate different aspects of the defensive responses. (C) 2008 Elsevier Inc. All rights reserved.

5-HT1A receptors of the lateral septum regulate inhibitory avoidance but not escape behavior in rats

Serotonin in the lateral septum (LS) has been implicated in the modulation of defensive behaviors and in anxiety. However, it is currently unknown whether changes in 5-HT mechanisms in this brain area may selectively affect defensive responses associated with specific subtypes of anxiety disorders recognized in clinical settings. To address this question, we evaluated the effect of the intra-LS injection of the 5-HT1A/7 receptor agonist 8-CH-DPAT (0.6, 3.0, 15.0 nmol) in male Wistar rats exposed to the elevated T-maze animal model of anxiety. This test allows the measurement of two behavioral defensive responses in the same rat: inhibitory avoidance and escape behavior. In clinical terms, these responses have been respectively related to generalized anxiety and panic disorder. The effects of 8-OH-DPAT were compared to those caused by a standard anxiolytic compound, the benzodiazepine receptor agonist midazolam (MDZ, 20 nmol). We also investigated whether the intra-LS injection of the 5-HT1A receptor antagonist WAY-100635 (0.37 nmol) was able to block the effects of 8-OH-DPAT. All animals were also tested in an open field for locomotor activity assessments. Results showed that whereas intra-LS administration of MDZ decreased avoidance latencies, suggesting an anxiolytic action, 8-OH-DPAT caused the opposite effect. Neither drug affected the escape performance. Intra-LS administration of WAY-100635 blocked the anxiogenic effect caused by 8-OH-DPAT. No changes to locomotion were detected in the open field. The data suggests that LS 5-HT1A receptors are involved in the control of inhibitory avoidance behavior and that a failure in this regulatory mechanism may be of importance to the physiopathology of generalized anxiety disorder. (c) 2008 Elsevier Inc. All rights reserved.

Antinociception induced by epidural motor cortex stimulation in naive conscious rats is mediated by the opioid system

Epidural motor cortex stimulation (MCS) has been used for treating patients with neuropathic pain resistant to other therapeutic approaches. Experimental evidence suggests that the motor cortex is also involved in the modulation of normal nociceptive response, but the underlying mechanisms of pain control have not been clarified yet. The aim of this study was to investigate the effects of epidural electrical MCS on the nociceptive threshold of naive rats. Electrodes were placed on epidural motor cortex, over the hind paw area, according to the functional mapping accomplished in this study. Nociceptive threshold and general activity were evaluated under 15-min electrical stimulating sessions. When rats were evaluated by the paw pressure test, MCS induced selective antinociception in the paw contralateral to the stimulated cortex, but no changes were noticed in the ipsilateral paw. When the nociceptive test was repeated 15 min after cessation of electrical stimulation, the nociceptive threshold returned to basal levels. On the other hand, no changes in the nociceptive threshold were observed in rats evaluated by the tail-flick test. Additionally, no behavioral or motor impairment were noticed in the course of stimulation session at the open-field test. Stimulation of posterior parietal or somatosensory cortices did not elicit any changes in the general activity or nociceptive response. Opioid receptors blockade by naloxone abolished the increase in nociceptive threshold induced by MCS. Data shown herein demonstrate that epidural electrical MCS elicits a substantial and selective antinociceptive effect, which is mediated by opioids. (C) 2008 Elsevier B.V. All rights reserved.

Cardiovascular responses to microinjection of noradrenaline into the medial amygdaloid nucleus of conscious rats result from alpha(2)-receptor activation and vasopressin release

The medial amygdaloid nucleus (MeA) is involved in the modulation of physiological and behavioral processes, as well as regulation of the autonomic nervous system. Moreover, MeA electrical stimulation evokes cardiovascular responses. Thus, as noradrenergic receptors are present in this structure, the present study tested the effects of local noradrenaline (NA) microinjection into the MeA on cardiovascular responses in conscious rats. Moreover, we describe the types of adrenoceptor involved and the peripheral mechanisms involved in the cardiovascular responses. Increasing doses of NA (3, 9, 27 or 45 nmol/100 nL) microinjected into the MeA of conscious rats caused dose-related pressor and bradycardic responses. The NA cardiovascular effects were abolished by local pretreatment of the MeA with 10 nmol/100 nL of the specific alpha(2)-receptor antagonist RX821002, but were not affected by local pretreatment with 10 nmol/100 nL of the specific alpha(1)-receptor antagonist WB4101. The magnitude of pressor response evoked by NA microinjected into the MeA was potentiated by intravenous pretreatment with the ganglion blocker pentolinium (5 mg/kg), and blocked by intravenous pretreatment with the selective V(1)-vasopressin antagonist dTyr(CH(2))(5)(Me)AVP (50 mu g/kg). In conclusion, our results show that microinjection of NA into the MeA of conscious rats activates local alpha(2)-adrenoceptors, evoking pressor and bradycardic responses, which are mediated by vasopressin release.

Involvement of the lateral habenula in the regulation of generalized anxiety- and panic-related defensive responses in rats

Recently obtained evidence points to the involvement of the lateral habenular nuclei (LHb) in the mediation of coping defensive responses to threatening/stressful stimuli. Nevertheless, the role of this brain area in the regulation of defensive responses that have been associated with specific subtypes of anxiety disorders recognized in clinical settings is presently unknown. To address this question, we investigated the effects of either electrolytic lesions or chemical stimulation of the LHb on the defensive behaviors generated in rats by the elevated T-maze. This experimental model allows the measurement, in a same rat, of two defensive behaviors, inhibitory avoidance and escape, that have been related in terms of psychopathology to generalized anxiety and panic disorders, respectively. Bilateral electrolytic lesions of the LHb (1 mA, 10 s) impaired inhibitory avoidance acquisition and facilitated escape performance. On the other hand, chemical stimulation of the LHb by bilateral microinjection of kainic acid (30-60 pmol/0.2 mu L) had the opposite effect, i.e., facilitated inhibitory avoidance and impaired escape. The present results indicate that the LHb exerts an opposed regulatory control on generalized anxiety- and panic-related defensive responses in rats. (c) 2008 Elsevier Inc. All rights reserved.

Cardiovascular effects of noradrenaline microinjection into the medial part of the superior colliculus of unanesthetized rats

The superior colliculus (SC) is a mesencephalic area involved in the mediation of defensive movements associated with cardiovascular changes. Noradrenaline (NA) is a neurotransmitter with an important role in central cardiovascular regulation exerted by several structures of the central nervous system. Although noradrenergic nerve terminals have been observed in the SC, there are no reports on the effects of local NA injection into this area. Taking this into consideration, we studied the cardiovascular effects of NA microinjection into the SC of unanesthetized rats. Microinjection of NA into the SC evoked a dose-dependent blood pressure increase and a heart rate decrease in unanesthetized rats. The pressor response to NA was not modified by intravenous pretreatment with the vasopressin v(1)-receptor antagonist dTyr(CH(2))(5) (Me)AVP, indicating a lack of vasopressin involvement in the response mediation. The effect of NA microinjection into the SC was blocked by intravenous pretreatment with the ganglionic blocker pentolinium, indicating its mediation by the sympathetic nervous system. Although the pressor response to NA was not affected by adrenal demedullation, the accompanying bradycardia was potentiated, suggesting some involvement of the sympathoadrenal system in the cardiovascular response to NA microinjection into the SC. In summary, results indicate that stimulation of noradrenergic receptors in the SC causes cardiovascular responses which are mediated by activation of both neural and adrenal sympathetic nervous system components. (C) 2009 Elsevier B.V. All rights reserved.

Oleoyl lysophosphatidic acid: a new mediator of emotional behavior in rats.

The role of lysophosphatidic acid (LPA) in the control of emotional behavior remains to be determined. We analyzed the effects of the central administration of 1-oleoyl-LPA (LPA 18∶1) in rats tested for food consumption and anxiety-like and depression-like behaviors. For this purpose, the elevated plus-maze, open field, Y maze, forced swimming and food intake tests were performed. In addition, c-Fos expression in the dorsal periaqueductal gray matter (DPAG) was also determined. The results revealed that the administration of LPA 18∶1 reduced the time in the open arms of the elevated plus-maze and induced hypolocomotion in the open field, suggesting an anxiogenic-like phenotype. Interestingly, these effects were present following LPA 18∶1 infusion under conditions of novelty but not under habituation conditions. In the forced swimming test, the administration of LPA 18∶1 dose-dependently increased depression-like behavior, as evaluated according to immobility time. LPA treatment induced no effects on feeding. However, the immunohistochemical analysis revealed that LPA 18∶1 increased c-Fos expression in the DPAG. The abundant expression of the LPA1 receptor, one of the main targets for LPA 18∶1, was detected in this brain area, which participates in the control of emotional behavior, using immunocytochemistry. These findings indicate that LPA is a relevant transmitter potentially involved in normal and pathological emotional responses, including anxiety and depression.

1-Oleoyl lysophosphatidic acid: a new mediator of emotional behavior in rats.

The role of lysophosphatidic acid (LPA) in the control of emotional behavior remains to be determined. We analyzed the effects of the central administration of 1-oleoyl-LPA (LPA 18∶1) in rats tested for food consumption and anxiety-like and depression-like behaviors. For this purpose, the elevated plus-maze, open field, Y maze, forced swimming and food intake tests were performed. In addition, c-Fos expression in the dorsal periaqueductal gray matter (DPAG) was also determined. The results revealed that the administration of LPA 18∶1 reduced the time in the open arms of the elevated plus-maze and induced hypolocomotion in the open field, suggesting an anxiogenic-like phenotype. Interestingly, these effects were present following LPA 18∶1 infusion under conditions of novelty but not under habituation conditions. In the forced swimming test, the administration of LPA 18∶1 dose-dependently increased depression-like behavior, as evaluated according to immobility time. LPA treatment induced no effects on feeding. However, the immunohistochemical analysis revealed that LPA 18∶1 increased c-Fos expression in the DPAG. The abundant expression of the LPA1 receptor, one of the main targets for LPA 18∶1, was detected in this brain area, which participates in the control of emotional behavior, using immunocytochemistry. These findings indicate that LPA is a relevant transmitter potentially involved in normal and pathological emotional responses, including anxiety and depression.

The indirect amygdala-dorsal striatum pathway mediates conditioned freezing: Insights on emotional memory networks

The dorsal striatum (DS) is involved in various forms of learning and memory such as procedural learning, habit learning, reward-association and emotional learning. We have previously reported that bilateral DS lesions disrupt tone fear conditioning (TFC), but not contextual fear conditioning (CFC) [Ferreira TL, Moreira KM, Ikeda DC, Bueno OFA, Oliveira MGM (2003) Effects of dorsal striatum lesions in tone fear conditioning and contextual fear conditioning. Brain Res 987:17-24]. To further elucidate the participation of DS in emotional learning, in the present study, we investigated the effects of bilateral pretest (postraining) electrolytic DS lesions on TFC. Given the well-acknowledged role of the amygdala in emotional learning, we also examined a possible cooperation between DS and the amygdala in TFC, by using asymmetrical electrolytic lesions, consisting of a unilateral lesion of the central amygdaloid nucleus (CeA) combined to a contralateral DS lesion. The results show that pre-test bilateral DS lesions disrupt TFC responses, suggesting that DS plays a role in the expression of TFC. More importantly, rats with asymmetrical pre-training lesions were impaired in TFC, but not in CFC tasks. This result was confirmed with muscimol asymmetrical microinjections in DS and CeA, which reversibly inactivate these structures. On the other hand, similar pretest lesions as well as unilateral electrolytic lesions of CeA and DS in the same hemisphere did not affect TFC. Possible anatomical substrates underlying the observed effects are proposed. Overall, the present results underscore that other routes, aside from the well-established CeA projections to the periaqueductal gray, may contribute to the acquisition/consolidation of the freezing response associated to a TFC task. It is suggested that CeA may presumably influence DS processing via a synaptic relay on dopaminergic neurons of the substantia nigra compacta and retrorubral nucleus. The present observations are also in line with other studies showing that TFC and CFC responses are mediated by different anatomical networks. (C) 2008 IBRO. Published by Elsevier Ltd. All rights reserved.

Anxiolytic-like effect of way-100635 microinfusions into the median (but not dorsal) raphe nucleus in mice exposed to the plus-maze: influence of prior test experience

Studies in several laboratories have confirmed the anxiolytic potential of a wide range of 5-HT1A receptor antagonists in rats and mice, with recent evidence pointing to a postsynaptic site of action in the ventral hippocampus. It would, therefore, be predicted that blockade of 5-HT1A somatodendritic autoreceptors in the midbrain raphe nuclei should produce anxiogenic-like effects. To test this hypothesis, we investigated the effects of WAY-100635 microinfusions (0, 1.0 or 3.0 mug in 0.1 mul) into the dorsal (DRN) or median (MRN) raphe nuclei on behaviours displayed by male Swiss-Webster mice in the elevated plus-maze. As this test is sensitive to prior experience. The effects of intra-raphe infusions were examined both in maze-naive and maze-experienced subjects. Sessions, were videotaped and subsequently scored for conventional indices of anxiety (open arm avoidance) and locomotor activity (closed arm entries), as well as a range of ethological measures (e.g. risk assessment). In maze-naive mice, intra-MRN (but not intra-DRN) infusions of WAY-100635 (3.0 mug) increased open arm exploration and reduced risk assessment. Importantly, these effects could not be attributed to a general reduction in locomotor activity. A similar, though somewhat weaker, pattern of behavioural change was observed in maze-experienced animals. This unexpected anxiolytic effect of 5-HT1A autoreceptor blockade in the MRN cannot be accounted fur by a disinhibition of 5-HT release in forebrain targets (e.g. hippocampus and amygdala), where stimulation of postsynaptic 5-HT1A receptors enhances anxiety-like responses. However, as the MRN also projects to the periaqueductal gray matter (PAG), an area known to be sensitive to the anti-aversive effects or 5-HT, it is argued that present results may reflect increased 5-HT release at this crucial midbrain locus within the neural circuitry of defense. (C) 2002 Elsevier B.V. B.V. All rights reserved.

The anatomical connections of the macaque monkey orbitofrontal cortex. A review

The orbitofrontal cortex (OfC) is a heterogeneous prefrontal sector selectively connected with a wide constellation of other prefrontal, limbic, sensory and premotor areas. Among the limbic cortical connections, the ones with the bippocampus and parabippocampal cortex are particularly salient. Sensory cortices connected with the OfC include areas involved in olfactory, gustatory, somatosensory, auditory and visual processing. Subcortical structures with prominent OfC connections include the amygdala, numerous thalamic nuclei, the striatum, hypothalamus, periaqueductal gray matter, and biochemically specific cell groups in the basal forebrain and brainstem. Architectonic and connectional evidence supports parcellation of the OfC. The rostrally placed isocortical sector is mainly connected with isocortical areas, including sensory areas of the auditory, somatic and visual modalities, whereas the caudal non-isocortical sector is principally connected with non-isocortical areas, and, in the sensory domain, with olfactory and gustatory areas. The connections of the isocortical and non- isocortical orbital sectors with the amygdala, thalamus, striatum, hypotbalamus and periaqueductal gray matter are also specific. The medial sector of the OfC is selectively connected with the bippocampus, posterior parabippocampal cortex, posterior cingulate and retrosplenial areas, and area prostriata, while the lateral orbitofrontal sector is the most heavily connected with sensory areas of the gustatory, somatic and visual modalities, with premotor regions, and with the amygdala.

Serotonergic modulation in neuropathy induced by oxaliplatin: Effect on the 5HT(2C) receptor

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

Neural correlates of sleepiness induced by catecholamine depletion

Although extensive indirect evidence exists to suggest that the central dopaminergic system plays a significant role in the modulation of arousal, the functional effect of the dopaminergic influence on the regulation of the sleep-wake cycle remains unclear. Thirteen healthy volunteers and 15 unmedicated subjects with a history of major depressive disorder underwent catecholamine depletion (CD) using oral alpha-methyl-para-tyrosine in a randomized, placebo-controlled, double-blind, crossover study. The main outcome measures in both sessions were sleepiness (Stanford-Sleepiness-Scale), cerebral glucose metabolism (positron emission tomography), and serum prolactin concentration. CD consistently induced clinically relevant sleepiness in both groups. The CD-induced prolactin increase significantly correlated with CD-induced sleepiness but not with CD-induced mood and anxiety symptoms. CD-induced sleepiness correlated with CD-induced increases in metabolism in the medial and orbital frontal cortex, bilateral superior temporal cortex, left insula, cingulate motor area and in the vicinity of the periaqueductal gray. This study suggests that the association between dopamine depletion and sleepiness is independent of the brain reward system and the risk for depression. The visceromotor system, the cingulate motor area, the periaqueductal gray and the caudal hypothalamus may mediate the impact of the dopaminergic system on regulation of wakefulness and sleep.

An investigation of the interrelationship between manipulative therapy-induced hypoalgesia and sympathoexcitation

Objective: To investigate a proposed model in which manipulative therapy produces a treatment-specific initial hypoalgesic and sympathoexcitatory effect by activating a descending pain inhibitory system. The a priori hypothesis tested was that manipulative therapy produces mechanical hypoalgesia and sympatho-excitation beyond that produced by placebo or control. Furthermore, these effects would be correlated, thus supporting the proposed model. Design: A randomized, double-blind, placebo-controlled, repeated-measures study of the initial effect of treatment. Setting: Clinical neurophysiology laboratory. Subjects: Twenty-four subjects (13 women and 11 men; mean age, 49 yr) with chronic lateral epicondylalgia (average duration, 6.2 months). Intervention: Cervical spine lateral glide oscillatory manipulation, placebo and control. Outcome Measures: Pressure pain threshold, thermal pain threshold, pain-free grip strength test, upper limb tension test 2b, skin conductance, pileous and glabrous skin temperature and blood flux. Results: Treatment produced hypoalgesic and sympathoexcitatory changes significantly grater than those of placebo and control (p < .03). Confirmatory factor-analysis modeling, which was performed on the pain-related measures and the indicators of sympathetic nervous system function, demonstrated a significant correlation (r = .82) between the latencies of manipulation-induced hypoalgesia and sympathoexcitation. The Lagrange Multiplier test and Wald test indicated that the two latent factors parsimoniously and appropriately represented their observed variables. Conclusions: Manual therapy produces a treatment-specific initial hypoalgesic and sympathoexcitatory effect beyond that of placebo or control. The strong correlation between hypoalgesic and sympathoexcitatory effects suggests that a central control mechanism might be activated by manipulative therapy.

The amygdaloid complex: Anatomy and physiology

A converging body of literature over the last 50 years has implicated the amygdala in assigning emotional significance or value to sensory information. In particular, the amygdala has been shown to be an essential component of the circuitry underlying fear-related responses. Disorders in the processing of fear-related information are likely to be the underlying cause of some anxiety disorders in humans such as posttraumatic stress. The amygdaloid complex is a group of more than 10 nuclei that are located in the midtemporal lobe. These nuclei can be distinguished both on cytoarchitectonic and connectional grounds. Anatomical tract tracing studies have shown that these nuclei have extensive intranuclear and internuclear connections. The afferent and efferent connections of the amygdala have also been mapped in detail, showing that the amygdaloid complex has extensive connections with cortical and subcortical regions. Analysis of fear conditioning in rats has suggested that long-term synaptic plasticity of inputs to the amygdala underlies the acquisition and perhaps storage of the fear memory. In agreement with this proposal, synaptic plasticity has been demonstrated at synapses in the amygdala in both in vitro and in vivo studies. In this review, we examine the anatomical and physiological substrates proposed to underlie amygdala function.