Fos-like immunoreactive neurons following electrical stimulation of the dorsal periaqueductal gray at freezing and escape thresholds

Electrical stimulation of the dorsal regions of the periaqueductal gray (PAG) leads to defensive reactions characterized as freezing and escape responses. Until recently it was thought that this freezing behavior could be due to the recruitment of neural circuits in the ventrolateral periaqueductal gray (vlPAG), while escape would be mediated by other pathways. Nowadays, this view has been changing mainly because of evidence that freezing and escape behaviors thus elicited are not altered after lesions of the vlPAG. It has been suggested that there are at least two pathways for periaqueductal gray-mediated defensive responses, one involving the hypothalamus and the cuneiform nucleus (CnF) which mediates responses to immediate danger and another one involving the amygdala and vlPAG which mediates cue-elicited responses, either learned or innate. To examine this issue further we measured Fos protein expression in brain areas activated by electrical stimulation of the dorsolateral PAG (dlPAG) at the freezing and escape thresholds. The data obtained showed that freezing-provoking stimulation caused increases in Fos expression in the dorsomedial PAG (dmPAG), while escape-provoking stimulation led to increases at both dmPAG and dlPAG. Surprisingly, neither escape- nor freezing-provoking stimulations altered Fos expression in the central nucleus of amygdala (CeA). Escape-provoking stimulation caused increased Fos expression in the ventromedial hypothalamus (VMH), dorsal premammilary nucleus (PMd) and in the cuneiform nucleus. Significant increases in Fos labeling were found in the dmPAG and PMd following freezing-provoking stimulation. Therefore, the present data support the notion of a neural segregation for defensive behaviors in the dorsal columns of PAG, with increased Fos expression in the dmPAG following freezing, while dlPAG is affected by both freezing and escape responses. dlPAG, CnF, VMH and PMd are part of a brain aversion network activated by fear unconditioned stimuli. The present data also suggests that the defensive responses generated at the dlPAG level do not recruit the neural circuits of the vlPAG and CeA usually activated by conditioned fear stimuli.     

Differential role of serotonin projections from the dorsal and median raphe nuclei in phencyclidine-induced hyperlocomotion and fos-like immunoreactivity in rats

Altered brain serotonin activity is implicated in schizophrenia. We have previously shown differential involvement of serotonergic projections from the dorsal or median raphe nucleus in phencyclidine‐induced hyperlocomotion in rats, a behavioral model of aspects of schizophrenia. Here we further investigated the effects of serotonergic lesions of the raphe nuclei on phencyclidine‐induced hyperlocomotion by parallel assessment of Fos‐like immunoreactivity (FLI), a marker of neuronal activation in the brain. Male Sprague‐Dawley rats were anesthetized with pentobarbitone and stereotaxically microinjected with 5 μg of the serotonergic neurotoxin, 5,7‐dihydroxytryptamine (5,7‐DHT), into either the dorsal raphe (DRN) or median raphe nucleus (MRN). Two weeks after the surgery, rats with lesions of the MRN, but not those with lesions of the DRN, showed significant enhancement of the hyperlocomotion induced by injection of 2.5 mg/kg of phencyclidine. Rats with MRN lesions also showed significantly higher levels of FLI in the polymorphic layer of the dentate gyrus in the dorsal hippocampus (PoDG) when compared with sham‐operated controls. Rats with lesions of the DRN showed significantly higher levels of FLI in the nucleus accumbens (NAcc). These results indicate that FLI in the PoDG, but not the NAcc, correlates with enhanced phencyclidine‐induced locomotor hyperactivity in MRN‐lesioned rats. These results support our previous studies suggesting a role of serotonergic projections from the MRN to the dorsal hippocampus in some of the symptoms of schizophrenia. Synapse, 2012. © 2012 Wiley Periodicals, Inc.     

Antidepressant treatment reduces Fos-like immunoreactivity induced by swim stress in different columns of the periaqueductal gray matter

Antidepressant treatment attenuates behavioral changes induced by uncontrollable stress. The periaqueductal gray matter (PAG) is proposed to be a brain site involved in the behavioral responses to uncontrollable stress and antidepressant effects. The main goal of the present study was to investigate the effect of antidepressant treatment on the pattern of neural activation of the PAG along its mediolateral and rostrocaudal subregions after a forced swim stress episode. Male Wistar rats were sub-acutely treated with desipramine (a selective noradrenaline re-uptake blocker, three injections of 10 mg/kg in 24 h) or clomipramine (a non-selective serotonin and noradrenaline re-uptake blocker, three injections of 10 mg/kg in 24 h) and submitted to the forced swimming test (FST). Two hours after the test their brain were removed for Fos immunohistochemistry. Fos-like immunoreactivity (FLI) in rostral, intermediate and caudal portions of dorsomedial (dmPAG), dorsolateral (dlPAG), lateral (lPAG) and ventrolateral (vlPAG) PAG were quantified by a computerized system. The FST session increased FLI in most parts of the PAG. Previous treatment with desipramine or clomipramine reduced FLI in all columns of the PAG. FLI in the PAG correlated positively with to the immobility time and negatively with to climbing behavior scored during the test. These results indicate that neurons in the PAG are activated by uncontrollable stress. Moreover, inhibitory action of antidepressants on this activity may be associated with the anti-immobility effects of these drugs in the FST.     

Fos expression induced by changes in arterial pressure is localized in distinct,longitudinally organized columns of neurons in the rat midbrain periaqueductal gray

The distribution of neurons expressing Fos within the periaqueductal gray (FAG) following pharmacologically induced high or low blood pressure was examined to determine (1) if PAG neurons are responsive to changes in arterial pressure (AP) and (2) the relationship of these cells to the functionally defined hypertensive and hypotensive columns in PAG. Changes in AP differentially induced robust Fos expression in neurons confined to discrete, longitudinally organized columns within PAG. Increased AP produced extensive Fos-like immunoreactivity within the lateral PAG, beginning at the level of the oculomotor nucleus. At the level of the dorsal raphe, Fos expression induced by increased AP shifted dorsally, into the dorsolateral division of PAG; this pattern of Fos labeling was maintained throughout the caudal one-third of PAG. Double-labeling for Fos and nicotinamide adenine dinucleotide phosphate diaphorase confirmed that Fos-positive cells induced by increased AP were located in the dorsolateral division of PAG at these caudal levels. Fos positive cells were codistributed, but not colocalized, with nicotinamide adenine dinucleotide phosphate diaphorase-positive cells. Decreased AP evoked a completely different pattern of Fos expression. Fos-positive cells were predominantly located within the ventrolateral PAG region, extending from the level of the trochlear nucleus through the level of the caudal dorsal raphe. Double-labeling studies for Fos and serotonin indicated that only 1–2 double-labeled cells per section were present. Saline infusion resulted in very few Foslike immunoreactive cells, indicating that volume receptor activation does not account for Fos expression in PAG evoked by changes in AP. These results indicate that (1) substantial numbers of PAG neurons are excited by pharmacologically induced changes in AP and (2) excitatory barosensitive PAG neurons are anatomically segregated based on their responsiveness to a specific directional change in AP. © 1995 Wiley-Liss, Inc.     

A reinvestigation of the analgesic effects induced by stimulation of the periaqueductal gray matter in the rat. I. The production of behavioral side effects together with analgesia

It has been shown that stimulation-produced-analgesia (SPA) in the cat elicited from the periaqueductal gray matter (PAG) is obtained from sites located in the ventral part, particularly the dorsal raphé nucleus (DRN). These data contrast with the numerous studies performed in the rat in which efficient sites seem widely distributed throughout the PAG. These discrepancies led us to reinvestigate SPA from PAG and adjacent structures in the rat. Central stimulation was delivered through bipolar concentric electrodes (one for each animal). Analgesia was evaluated (before and during central stimulation) by measuring the modification in the vocalization threshold induced by electrical tail shocks or by considering the reaction of the animal to pinch. In contrast with the majority of previous studies, these experiments were performed on the totally freely-moving rat. The most striking result was that, in order to obtain analgesia from all regions of the PAG, it was necessary to apply intensities of central stimulation which also triggered other strong behavioral reactions. With intensities of PAG stimulation which did not induce such side effects, very few effective analgesic sites were found (21/129 sites of which 14/83 were strictly located in the PAG). However, it was possible to define two 'pure analgesic regions', both located in the ventral PAG: one centered on the dorsomedial part of the DRN and the other one situated in the ventrolateral PAG. No modification of nociceptive thresholds was observed when stimulating the dorsal and dorsolateral parts of the PAG as well as structures adjacent to these regions; in some rats, an increase in pain reactivity was even noted. When the intensity of central stimulation (applied to the various parts of the PAG) was increased, some stereotyped 'behavioral responses' occurred depending on the location of the stimulation site: motor effects (gnawing, rotation or tremor) in the ventral PAG and aversive effects (flight, jumping and on occasions, distress vocalizations) in the dorsal, dorsolateral PAG and in the ventral region just surrounding the cerebral aqueduct. Under these conditions, analgesia was obtained from practically the entire PAG, the vocalization threshold being increased dramatically on occasions. It must be emphasized that antinociceptive effects associated with other obvious behavioral manifestations (aversive ones) were also obtained from sites located outside the PAG (colliculi and tectum adjacent to the dorsal and dorsolateral PAG).(ABSTRACT TRUNCATED AT 400 WORDS)     

Neural circuitry controlling vasopressin-stimulated scent marking in Syrian hamsters (Mesocricetus auratus)

In Syrian hamsters, arginine vasopressin (AVP) plays a critical role in the control of a form of scent marking called flank marking. Microinjection of AVP into the medial preoptic-anterior hypothalamus (MPOA-AH), lateral septal nucleus (LS), bed nucleus of the stria terminalis (BNST), and the periaqueductal gray (PAG) stimulates high levels of flank marking. Microinjection of an antagonist of the V_1a-AVP receptor into sites such as the MPOA-AH inhibits expression of flank marking. The purpose of the present study was to investigate the neural circuit controlling flank marking by localizing the induction of Fos protein in response to the microinjection of AVP, a V_1a-AVP antagonist (AVPA) or saline into the MPOA-AH. Immediately after microinjection, hamsters were placed in a clean cage and their behavior was videotaped for 10 minutes. Ninety minutes after the behavioral experiment hamsters were perfused and their brains removed for subsequent immunocytochemical localization of Fos protein. The number of Fos-positive neurons was significantly greater in the BNST, PAG, and central amygdala (Ce) following the microinjection of AVP than following the microinjection of either AVPA or saline. In AVP-injected animals, the number of Fos-labeled cells in the Ce, PVN, and PAG increased with increased frequency of either flank marking or flank gland grooming. These data support the hypothesis that neurons within the MPOA-AH, BNST, and PAG play an important role in the control of flank marking and suggest that the Ce may be a previously unrecognized part of this neural circuit. © 1996 Wiley-Liss, Inc.     

The efferent projections of the periaqueductal gray in the rat: A Phaseolus vulgaris-leucoagglutinin study. II. Descending projections

The descending projections of the periaqueductal gray (PAG) have been studied in the rat using the anterograde tracer Phaseolus vulgaris-leucoagglutinin. The tracer was injected into the dorsolateral or ventrolateral subdivisions of the PAG at rostral or caudal sites. It was found that the patterns of the descending projections of the rostral and caudal parts of the dorsolateral PAG were the same and that the patterns of the descending projections of the rostral and caudal parts of the ventrolateral PAG were the same. However, the patterns of projections of the dorsolateral and ventrolateral PAG subregions were substantially different. These results suggest that the dorsolateral and ventrolateral parts of the PAG are organized into longitudinal columns that extend throughout the length of the PAG. The axons of PAG neurons descended through the pons and medulla via two routes. A small fiber bundle was present in the periaqueductal gray and in the periventricular area. This bundle distributed fibers and terminals locally within the periaqueductal gray and in the locus coeruleus and Barrington's nucleus. A larger bundle had a diffuse arrangement in the pontine reticular formation, however, and it had a more restricted distribution in the medulla, where it occupied a position dorsolateral to the pyramid. This bundle supplied structures in the pontine and medullary tegmentum. The dorsolateral column preferentially supplied the locus coeruleus, subcoeruleus, the gigantocellular nucleus pars alpha, the rostral part of the paragigantocellular nucleus, and the region of the A5 noradrenergic cell group. The ventrolateral column preferentially supplied the nucleus raphe magnus, the caudal part of the lateral paragigantocellular nucleus, and the rostroventrolateral reticular nucleus. © 1995 Willy-Liss, Inc.     

Predatory hunting and exposure to a live predator induce opposite patterns of Fos immunoreactivity in the PAG

Considering the periaqueductal gray's (PAG) general roles in mediating motivational responses, in the present study, we compared the Fos expression pattern in the PAG induced by innate behaviors underlain by opposite motivational drivers, in rats, namely, insect predation and defensive behavior evoked by the confrontation with a live predator (a cat). Exposure to the predator was associated with a striking Fos expression in the PAG, where, at rostral levels, an intense Fos expression was found largely distributed in the dorsomedial and dorsolateral regions, whereas, at caudal levels, Fos-labeled cells tended to be mostly found in the lateral and ventrolateral columns, as well as in the dorsal raphe nucleus. Quite the opposite, insect predation was associated with increased Fos expression predominantly in the rostral two thirds of the lateral PAG, where the majority of the Fos-immunoreactive cells were found at the oculomotor nucleus levels. Remarkably, both exposure to the cat and insect predation upregulated Fos expression in the supraoculomotor region and the laterodorsal tegmental nucleus. Overall, the present results clearly suggest that the PAG activation pattern appears to reflect, at least partly, the animal's motivational status. It is well established that the PAG is critical for the expression of defensive responses, and, considering the present findings, it will be important to investigate how the PAG contributes to the expression of the predatory behavior, as well.     

Acoustic brainstem nuclei express Fos after flurothyl-induced generalized seizures in rats

The inferior colliculus (IC) plays a key role in modulating audiogenic seizures (AS) in rats. We investigated whether acoustic brainstem nuclei express Fos-like immunoreactivity (FLI) after flurothyl-induced generalized seizures in rats. Compared to controls, experimental animals showed significantly (P<0.05) more FLI in the dorsal and external cortex of the IC, as well as in the medial part of the medial geniculate body (MGB), perigeniculate area, and dorsal cochlear nucleus. No significant increase of FLI was observed in the central nucleus of the IC, ventral and dorsal parts of the MGB, dorsal nucleus of the lateral lemniscus, or ventral cochlear nucleus. Because this pattern of FLI closely resembles that observed after AS in previous studies, these results suggest that Fos expression in acoustic brainstem nuclei is not specific for AS.     

Fos-like immunoreactivity in the periaqueductal gray of rats exposed to a natural predator

In the present study we examined, in rats exposed to a predator (cat), the distribution of neurons expressing Fos along the continuum formed by the central gray surrounding the caudal pole of the third ventricle and the periaqueductal gray (PAG). After the predatory encounter, a distinct cluster of Fos-immunoreactive cells was observed in the precommissural nucleus. In the rostral two-thirds of the PAG, Fos expression was mostly seen in the dorsomedial and dorsolateral regions. In contrast, at caudal levels of the PAG, most of the Fos-labelled neurons were distributed in the lateral and ventrolateral PAG. These results are discussed and compared with the pattern of the PAG activation after fear conditioned to a context or elicited by aversive foot shock.     

Subregions of the periaqueductal gray topographically innervate the rostral ventral medulla in the rat.

Previous anatomical and physiological studies have revealed a substantial projection from the periaqueductal gray (PAG) to the nucleus paragigantocellularis (PGi). In addition, physiological studies have indicated that the PAG is composed of functionally distinct subregions. However, projections from PAG subregions to PGi have not been comprehensively examined. In the present study, we sought to examine possible topographic specificity for projections from subregions of the PAG to PGi. Pressure or iontophoretic injections of wheat germ agglutinin-conjugated horseradish peroxidase, or of Fluoro-Gold, placed into the PGi of the rat retrogradely labeled a substantial number of neurons in the PAG from the level of the Edinger-Westphal nucleus to the caudal midbrain. Retrogradely labeled neurons were preferentially aggregated in distinct subregions of the PAG. Rostrally, at the level of the oculomotor nucleus, labeled neurons were i) compactly aggregated in the ventromedial portion of the PAG corresponding closely to the supraoculomotor nucleus of the central gray, ii) in the lateral and ventrolateral PAG, and iii) in medial dorsal PAG. More caudally, retrogradely labeled neurons became less numerous in the dorsomedial PAG but were more widely scattered throughout the lateral and ventrolateral parts of the PAG. Only few retrogradely labeled neurons were found in the ventromedial part of the PAG at caudal levels. Injections of retrograde tracers restricted to subregions of the PGi suggested topography for afferents from the PAG. Injections into the lateral portion of the PGi yielded the greatest number of labeled neurons within the rostral ventromedial PAG. Medially placed injections yielded numerous retrogradely labeled neurons in the lateral and ventrolateral PAG. Injections placed in the rostral pole of the PGi (medial to the facial nucleus) produced the greatest number of retrogradely labeled neurons in the dorsal PAG. To examine the pathways taken by fibers projecting from PAG neurons to the medulla, and to further specify the topography for the terminations of these afferents in the PGi, the anterograde tracer Phaseolus vulgaris-leucoagglutinin was iontophoretically deposited into subregions of the PAG that contained retrogradely labeled neurons in the above experiments. These results revealed distinct fiber pathways to the rostral medulla that arise from the dorsal, lateral/ventrolateral, and ventromedial parts of the PAG. These injections also showed that there are differential but overlapping innervation patterns within the PGi. Consistent with the retrograde tracing results, injections into the rostral ventromedial PAG near the supraoculomotor nucleus yielded anterograde labeling immediately ventral to the nucleus ambiguus in the ventrolateral medulla, within the retrofacial portion of the PGi.(ABSTRACT TRUNCATED AT 400 WORDS)     

Effects of 21 days treatment with fluoxetine on stimulated endogenous 5-hydroxytryptamine overflow in the rat dorsal raphe and suprachiasmatic nucleus studies using fast cyclic voltammetry in vitro

Changes in the extracellular concentration of 5-HT evoked by electrical stimulation of brain slices containing either dorsal raphe nucleus (DRN) or suprachiasmatic nucleus (SCN) from rats treated for 21 days with fluoxetine (5 mg/kg; i.p.) or water were monitored using fast cyclic voltammetry (FCV). Stimulated 5-HT overflow was enhanced significantly in both brain regions after 21 days treatment with fluoxetine but there was no change in the half time for re-uptake (t1/2). Concentration response curves for inhibition of electrically stimulated 5-HT overflow by 8-OH-DPAT (5-HT_1a receptor agonist) or RU24969 (5-HT_1b receptor agonist) in the DRN or SCN respectively were obtained in slices prepared from both groups of animals. There was a significant shift to the right in the dose-response curve for RU24969 in the SCN in fluoxetine treated animals but a shift to the left for the dose-response curve for 8-OH-DPAT in the DRN. These data suggest that down regulation of the 5-HT_1b autoreceptors occurs in an axon terminal region (SCN) but that there is a sensitisation of 5-HT_1a autoreceptor mechanisms controlling 5-HT overflow in the DRN.     

Increased c-Fos expression in select lateral parabrachial subnuclei following chemical versus electrical stimulation of the dorsal periaqueductal gray in rats

The parabrachial nucleus (PBN) is located in the rostral dorsolateral pons and has been identified as a critical relay for cardiovascular responses (sympathoexcitation and baroreflex attenuation) evoked by the dorsal periaqueductal gray (PAG). We examined the pattern of c-Fos protein immunoreactivity throughout the rostral-caudal extent of the PBN in four groups of anesthetized male Sprague-Dawley rats to identify the specific PBN regions activated by dorsal PAG stimulation. Both electrical stimulation and chemical (0.3 mM bicuculline methobromide) activation of the dorsal PAG elicited a selective increase in Fos-like immunoreactivity (FLI) in the superior lateral and central lateral subnuclei of the rostral lateral PBN (LPBN) relative to surgery and blood pressure control groups. In the middle LPBN chemical stimulation of the dorsal PAG selectively increased FLI in the central lateral subnucleus while electrical stimulation increased FLI in the Kolliker-Fuse area only. Finally, in the caudal LPBN only electrical stimulation of the dorsal PAG induced significant changes in FLI above control. Significant changes in FLI in the medial PBN were not observed under any experimental conditions. These results confirm neuroanatomical data demonstrating that neurons in superior lateral and central lateral subnuclei of the rostral and middle LPBN are the primary targets of the dorsal PAG. Our results also demonstrate that this descending projection to the central lateral and superior lateral subnuclei of the LPBN is in part excitatory. Finally, our results raise the possibility that neurons in the central lateral subnucleus of the middle and rostral LPBN are integrally involved in descending modulation of sympathetic drive associated with dorsal PAG activation.     

The consequences of uncontrollable stress are sensitive to duration of prior wheel running

The behavioral consequences of uncontrollable stress, or learned helplessness (LH) behaviors, are thought to involve hyperactivity of serotonergic (5-HT) neurons in the dorsal raphe nucleus (DRN). Other brain regions implicated in LH and capable of affecting 5-HT systems, such as the bed nucleus of the stria terminalis (BNST), amygdala, and habenula, could contribute to DRN 5-HT hyperactivity during uncontrollable stress. Six weeks of wheel running prevents LH and attenuates uncontrollable stress-induced c-Fos expression in DRN 5-HT neurons, although the duration of wheel running necessary for these effects is unknown. In the current study, 6 but not 3, weeks of wheel running blocked the shuttle box escape deficit and exaggerated fear produced by uncontrollable tail shock in sedentary rats. Corresponding to the duration-dependent effects of wheel running on LH behaviors, 6 weeks of wheel running was required to attenuate uncontrollable stress-induced 5-HT neural activity, indexed by c-Fos protein expression, in the DRN and c-Fos expression in the lateral ventral region of the BNST. Wheel running, regardless of duration, did not affect c-Fos expression anywhere in the amygdala or habenula. These data indicate that the behavioral effects of uncontrollable stress are sensitive to the duration of prior physical activity and are consistent with the hypothesis that attenuation of DRN 5-HT activity contributes to the prevention of LH by wheel running. The potential role of the BNST in the prevention of LH by wheel running is discussed.     

Fos expression induced by warming the preoptic area in rats

The preoptic area (POA) occupies a crucial position among the structures participating in thermoregulation, but we know little about its efferent projections for controlling various effector responses. In this study, we used an immunohistochemical analysis of Fos expression during local warming of the preoptic area. To avoid the effects of anesthesia or stress, which are known to elicit Fos induction in various brain regions, we used a novel thermode specifically designed for chronic warming of discrete brain structures in freely moving rats. At an ambient temperature of 22 degrees C, local POA warming increased Fos immunoreactivity in the supraoptic nucleus (SON) and the periaqueductal gray matter (PAG). Exposure of animals to an ambient temperature of 5 degrees C induced Fos immunoreactivity in the magnocellular paraventricular nucleus (mPVN) and the dorsomedial region of the hypothalamus (DMH). Concurrent warming of the POA suppressed Fos expression in these areas. These findings suggest that thermal information from the preoptic area sends excitatory signals to the SON and the PAG, and inhibitory signals to the mPVN and the DMH.     

A critical role for the parabrachial nucleus in generating central nervous system responses elicited by a systemic immune challenge

Using Fos immunolabelling as a marker of neuronal activation, we investigated the role of the parabrachial nucleus in generating central neuronal responses to the systemic administration of the proinflammatory cytokine interleukin-1beta (1 microg/kg, i.a.). Relative to intact animals, parabrachial nucleus lesions significantly reduced the number of Fos-positive cells observed in the central amygdala (CeA), the bed nucleus of the stria terminalis (BNST), and the ventrolateral medulla (VLM) after systemic interleukin-1beta. In a subsequent experiment in which animals received parabrachial-directed deposits of a retrograde tracer, it was found that many neurons located in the nucleus tractus solitarius (NTS) and the VLM neurons were both retrogradely labelled and Fos-positive after interleukin-1beta administration. These results suggest that the parabrachial nucleus plays a critical role in interleukin-1beta-induced Fos expression in CeA, BNST and VLM neurons and that neurons of the NTS and VLM may serve to trigger or at least influence changes in parabrachial nucleus activity that follows systemic interleukin-1beta administration.     

The comparative effects of environmental enrichment with exercise and serotonin transporter blockade on serotonergic neurons in the dorsal raphe nucleus

We have previously reported that inhibition of the serotonin transporter (SERT) by selective serotonin reuptake inhibitor (SSRI) fluoxetine significantly reduces the number of tryptophan hydroxylase (TPH)-positive cells in the dorsal raphe nucleus (DRN). We have been interested in exploring whether this SSRI-induced change in TPH might be modified by housing in an enriched environment. Like SSRI antidepressants, environmental enrichment (EE) and physical exercise have been found to have efficacy in the prevention and alleviation of depression. We postulated that EE with exercise and SERT inhibition would similarly affect TPH regulation and that EE with exercise might modify the effect of fluoxetine on TPH. Three week old male Sprague-Dawley rats were housed in either a standard cage (SE) or an enriched environment (EE). SE animals were singly housed with no access to enrichment objects. EE animals were group housed and were provided with various enrichment objects (e.g. running wheel) that were changed and rearranged regularly. Nine weeks after the experiment began, the rats were randomly assigned to one of four treatment groups: (1) SE control; (2) SE fluoxetine; (3) EE control; or (4) EE fluoxetine. Fluoxetine (5 mg/kg/day) was placed in the drinking water. Sections of DRN were processed for TPH immunohistochemistry. The number of TPH-positive cells was determined by blinded, manual counting. Results were analyzed by analysis of variance (ANOVA) followed by post-hoc Tukey tests. Significance was set at P < 0.05. For animals housed in a standard environment, fluoxetine induced a significant 29% reduction in the number of TPH-immunoreactive cells in the DRN. A similar reduction in TPH immunoreactivity was observed in animals that were housed in an enriched environment but not exposed to fluoxetine (39%). The number of TPH-positive cells in the DRN for animals housed in an enriched environment and exposed to fluoxetine was not significantly different than animals housed in an enriched environment and not exposed to fluoxetine. The reduction of TPH immunoreactivity in the DRN by EE with exercise suggests that a modified housing environment and voluntary exercise affects regulation of TPH, possibly via a mechanism similar to that of SERT inhibitors. This downregulation of serotonin biosynthesis by fluoxetine and EE with exercise may ultimately play a role in the therapeutic action of both interventions.     

N-acetyl-aspartylglutamate: binding sites and excitatory action in the dorsolateral septum of rats

In this study we examined the distribution of binding sites for [3H]N-acetyl-aspartylglutamate (NAAG) in the rat lateral septal nucleus (LSN) and the effect of iontophoretically applied NAAG on neuronal firing in this area. A high density of [3H]NAAG binding sites was found in the dorsolateral part of the LSN. Binding in the intermediate/ventral part of the LSN and medial septum was less dense. NAAG excited 75% of the dorsal neurons in the LSN, but only 36% of the cells in the intermediate/ventral part. Glutamic diethylester, an amino acid antagonist, depressed responses to NAAG to a similar extent as responses to quisqualate. The antagonist amino phosphonovaleric acid, which suppressed responses to N-methyl-D-aspartate almost completely, reduced NAAG-evoked responses only by 40%. A possible role of NAAG as excitatory transmitter in the LSN is discussed.     

Hypothalamic paraventricular nucleus neurons regulate medullary catecholamine cell responses to restraint stress

Both physical and psychological stressors recruit catecholamine cells (CA) located in the ventrolateral medulla (VLM) and the nucleus of the solitary tract (NTS). In the case of physical stressors, this effect is initiated by signals that first access the central nervous system at or below the level of the medulla. For psychological stressors, however, CA cell recruitment depends on higher structures within the neuraxis. Indeed, we have recently provided evidence of a pivotal role for the medial amygdala (MeA) in this regard, although such a role must involve a relay, as MeA neurons do not project directly to the medulla. However, some of the MeA neurons that respond to psychological stress have been found to project to the hypothalamic paraventricular nucleus (PVN), a structure that provides significant input to the medulla. To determine whether the PVN might regulate medullary CA cell responses to psychological stress, animals were prepared with unilateral injections of the neurotoxin ibotenic acid into the PVN (Experiment 1), or with unilateral injections of the retrograde tracer wheat germ agglutinin-gold (WGA-Au) into the CA cell columns of the VLM or NTS (Experiment 2). Seven days later, animals were subjected to a psychological stressor (restraint; 15 minutes), and their brains were subsequently processed for Fos plus appropriate cytoplasmic markers (Experiment 1), or Fos plus WGA-Au (Experiment 2). PVN lesions significantly suppressed the stress-related induction of Fos in both VLM and NTS CA cells, whereas tracer deposits in the VLM or NTS retrogradely labeled substantial numbers of PVN cells that were also Fos-positive after stress. Considered in concert with previous results, these data suggest that the activation of medullary CA cells in response to psychological stress may involve a critical input from the PVN.