Seizure frequency reduction after posteromedial hypothalamus deep brain stimulation in drug-resistant epilepsy associated with intractable aggressive behavior.

OBJECTIVES: The aim of this study was to analyze the impact of deep brain stimulation (DBS) of the posteromedial hypothalamus (pHyp) on seizure frequency in patients with drug-resistant epilepsy (DRE) associated with intractable aggressive behavior (IAB). METHODS: Data were collected retrospectively from nine patients, who received bilateral stereotactic pHyp-DBS for the treatment of medically intractable aggressive behavior, focusing on five patients who also had DRE. All patients were treated at the Colombian Center and Foundation of Epilepsy and Neurological Diseases-FIRE (Chapter of the International Bureau for Epilepsy), in Cartagena de Indias, Colombia from 2010 to 2014. Each case was evaluated previously by the institutional ethical committee, assessing the impact of aggressive behavior on the patient's family and social life, the humanitarian aspects of preserving the safety and physical integrity of caregivers, and the need to prevent self-harm. Epilepsy improvement was measured by a monthly seizure reduction percentage, comparing preoperative state and outcome. Additional response to epilepsy was defined by reduction of the antiepileptic drugs (AEDs). Aggressive behavior response was measured using the Overt Aggression Scale (OAS). RESULTS: All the patients with DRE associated with IAB presented a significant decrease of the rate of epileptic seizures after up to 4 years follow-up, achieving a general 89.6% average seizure reduction from the state before the surgery. Aggressiveness was significantly controlled, with evident improvement in the OAS, enhancing the quality of life of patients and families. SIGNIFICANCE: In well-selected patients, DBS of the pHyp seems to be a safe and effective procedure for treatment of DRE associated with refractory aggressive behavior. Larger and prospective series are needed to define the pHyp as a target for DRE in different contexts.

Fatigue is mediated by cholinoceptors within the ventromedial hypothalamus independent of changes in core temperature.

We investigated brain mechanisms modulating fatigue during prolonged physical exercise in cold environments. In a first set of studies, each rat was subjected to three running trials in different ambient temperatures (T(a)). At 8 °C and 15 °C, core body temperature (T(core)) decreased and increased, respectively, whereas at 12 °C, the T(core) did not change throughout the exercise. In another set of experiments, rats were randomly assigned to receive bilateral 0.2 µL injections of 2.5 × 10(-2) M methylatropine or 0.15 M NaCl solution into the ventromedial hypothalamic nuclei (VMH). Immediately after the injections, treadmill exercise was started. Each animal was subjected to two experimental trials at one of the following T(a) : 5 °C, 12 °C or 15 °C. Muscarinic blockade of the VMH reduced the time to fatigue (TF) in cold environments by 35-37%. In all T(a) studied, methylatropine-treated rats did not present alterations in T(core) and tail skin temperature compared with controls. These results indicate that, below the zone of thermoneutrality, muscarinic blockade of the VMH decreases the TF, independent of changes in T(core). In conclusion, our data suggest that VMH muscarinic transmission modulates physical performance, even when the effects of thermoregulatory adjustments on fatigue are minimal.

Neuropeptide Y in the medial basal hypothalamus and medial preoptic area during the induction of LH surge may be controlled by locus coeruleus.

The multiple control of gonadotropin releasing hormone (GnRH)/luteinizing hormone (LH) secretion involves locus coeruleus (LC) and neuropeptide Y (NPY). The objective of the present study was to analyze the possible contribution of the LC to the control of NPY activity in the medial basal hypothalamus (MBH) and medial preoptic area (MPOA) during the LH surge induced by estrogen (E(2)) and progesterone (P(4)). Ovariectomized adult Wistar rats were submitted to the hormone replacement and to the LC bilateral lesion (lesioned groups) or sham surgery (control groups). On the day of the experiment the rats were decapitated at 11:00, 13:00, 15:00 and 17:00 h for plasma and brain collection. Plasma LH was determined by radioimmunoassay. MBH and MPOA were microdissected for the measurement of NPY by enzyme immunoassay. NPY mRNA levels in MBH were assessed by the ribonuclease protection assay. The results showed that LC lesion: decreased the plasma LH; increased the content of NPY in the MBH and reduced the increase of NPY content in the MPOA during afternoon in which LH surge was induced. The increased NPY content in MBH was not associated with an increase of the respective mRNA content, suggesting the action of postranscriptional and/or postranslational mechanisms. In conclusion, the NPY activity in the MPOA on LH surge induced by estrogen and progesterone could be controlled by LC through two ways, at least: one direct way, by the release of NPY from LC neurons terminals that innervate the MPOA and they release NA and NPY; one indirect way, by the control of release but not synthesis of NPY from neurons in the MBH which innervate the MPOA.

Hypothalamic tanycytes: a key component of brain-endocrine interaction.

Tanycytes are bipolar cells bridging the cerebrospinal fluid (CSF) to the portal capillaries and may link the CSF to neuroendocrine events. During the perinatal period a subpopulation of radial glial cells differentiates into tanycytes, a cell lineage sharing some properties with astrocytes and the radial glia, but displaying unique and distinct morphological, molecular, and functional characteristics. Four populations of tanycytes, alpha(1,2) and beta(1,2), can be distinguished. These subtypes express differentially important functional molecules, such as glucose and glutamate transporters; a series of receptors for neuropeptide and peripheral hormones; secretory molecules such as transforming growth factors, prostaglandin E(2), and the specific protein P85; and proteins of the endocytic pathways. This results in functional differences between the four subtypes of tanycytes. Thus, alpha(1,2) tanycytes do not have barrier properties, whereas beta(1,2) tanycytes do. Different types of tanycytes use different mechanisms to internalize and transport cargo molecules; compounds internalized via a clathrin-dependent endocytosis would only enter tanycytes from the CSF. There are also differences in the neuron-tanycyte relationships; beta(1,2) tanycytes are innervated by peptidergic and aminergic neurons, but alpha(1,2) tanycytes are not. Important aspects of the neuron-beta(1) tanycyte relationships have been elucidated. Tanycytes can participate in the release of gonadotropin-releasing hormone (GnRH) to the portal blood by expressing estrogen receptors, absorbing molecules from the CSF, and providing signal(s) to the GnRH neurons. Removal of tanycytes prevents the pulse of GnRH release into the portal blood, the peak of luteinizing hormone, and ovulation. The discovery in tanycytes of new functional molecules is opening a new field of research. Thus, thyroxine deiodinase type II, an enzyme generating triiodothyronine (T(3)) from thyroxine, appears to be exclusively expressed by tanycytes, suggesting that these cells are the main source of brain T(3). Glucose transporter-2 (GLUT-2), a low-affinity transporter of glucose and fructose, and ATP-sensitive K(+) channels are expressed by tanycytes, suggesting that they may sense CSF glucose concentrations.

Lateral hypothalamus lesions influences water and salt intake, and sodium and urine excretion, and arterial blood pressure induced by L-NAME and FK 409 injections into median preoptic nucleus in conscious rats.

Male Holtzman rats weighting 200-250 g were anesthetized with zoletil 50 mg/Kg (tiletamine chloridrate 125,0 mg and zolazepan chloridrate 125,0 mg) into quadriceps muscle and submitted an electrolytic lesion of the lateral hypothalamus (LH) and a stainless steel cannula was implanted into their median preoptic nucleus (MnPO). We investigated the effects of the injection into the (MnPO) of FK 409 (20 microg/0.5 microl), a nitric oxide (NO) donor, and N(W)-nitro-L-arginine methyl ester (L-NAME) 40 microg/0.5 microl, a nitric oxide synthase inhibitor (NOSI), on the water and sodium appetite and the natriuretic, diuretic and cardiovascular effects induced by injection of L-NAME and FK 409 injected into MnPO in rats with LH lesions. Controls were injected with a similar volume of 0.15 M NaCl. L-NAME injected into MnPO produced an increase in water and sodium intake and in sodium and urine excretion and increase de mean arterial pressure (MAP). FK 409 injected into MnPO did not produce any change in the hydro electrolytic and cardiovascular parameters in LH-sham and lesioned rats. FK 409 injected before L-NAME attenuated its effects. These data show that electrolytic lesion of the LH reduces fluid and sodium intake as well as sodium and urine excretion, and the pressor effect induced by L-NAME. LH involvement with NO of the MnPO excitatory and inhibitory mechanisms related to water and sodium intake, sodium excretion and cardiovascular control is suggested.

Organizaçäo neural de diferentes tipos de medo e suas implicaçöes na ansiedade / Neural organization of different types of fear: implications for the understanding of anxiety

A natureza das respostas de medo em animais expostos a situaçöes ameaçadoras depende da intensidade e da distância do estímulo aversivo. Esses estímulos podem ser potencialmente perigosos, distais ou proximais ao animal. Esforços têm sido feitos no sentido de identificar os circuitos neurais recrutados na organizaçäo das reaçöes defensivas a estas condiçöes aversivas. Neste artigo, sumarizamos evidências que associam os sistemas cerebrais de defesa ao conceito de medo-stress-ansiedade. Respostas de orientaçäo ao estímulo de perigo, à esquiva e à preparaçäo para o enfrentamento do perigo parecem estar associados à ansiedade. O giro do cíngulo e o córtex pré-frontal de um lado; o núcleo mediano da rafe, septo e o hipocampo de outro fazem parte dos circuitos cerebrais que integram essas respostas emocionais. No outro extremo, estímulos de medo que induzem formas ativas de defesa, mas pouco elaboradas, determinam estados emocionais de natureza diferente e parecem associadas a manifestaçöes elementares de medo. A substância cinzenta periaquedutal dorsal constitui o principal substrato neural para a integraçäo desses estados aversivos no cérebro. Comportamentos defensivos desse tipo säo produzidos pela estimulaçäo elétrica e química desta estrutura. A medida que os estímulos ameaçadores, potenciais e distais däo lugar a estímulos de perigo muito intensos ou säo substituídos por estímulos proximais de medo, ocorre uma comutaçäo (switch) dos circuitos neurais usualmente responsáveis pela produçäo de respostas condicionadas de medo para reaçöes defensivas com baixo nível de regulaçäo e organizaçäo que se assemelham aos ataques de pânico. Portanto, dependendo da natureza do evento estressor ou do estímulo incondicionado, o padräo de respostas defensivas orientadas e organizadas cede lugar a respostas motoras incoordenadas e incompletas. A amígdala e o hipotálamo medial podem funcionar como uma espécie de interface comutando os estímulos para os substratos neurais apropriados para elaboraçäo das respostas defensivas condicionadas ou incondicionadas

The medial hypothalamic defensive system: hodological organization and functional implications.

The hypothalamus is a relatively small division of the vertebrate forebrain that plays especially important roles in neural mechanisms assuring homeostasis, defense, and reproduction. Previous studies from our laboratory have suggested a distinct circuit in the medial hypothalamic zone as critically involved in the organization of innate defensive behavior. Thus, after exposure to a natural predator known to elicit innate defensive responses, increased Fos levels in the medial zone of the hypothalamus have been found restricted to the anterior hypothalamic nucleus, dorsomedial part of the ventromedial nucleus, and dorsal premammillary nucleus (PMd). Previous anatomical studies have shown that these Fos-responsive cell groups in the medial hypothalamus are interconnected in a distinct neural system, in which the PMd appears to be a critical element for the expression of defensive responses elicited by the presence of a predator. The purpose of this review is to provide an overview of what is currently known about the functional and hodological organization of this hypothalamic circuit subserving defensive responses.

Participation of the medial and anterior hypothalamus in the modulation of tonic immobility in guinea pigs.

Tonic immobility (TI) is an inhibitory behavioral response during which the animal presents profound physical inactivity and a relative lack of response to the environment. This response is induced in the laboratory by postural inversion of the animal and brief postural contention of its movements. In nature, the response occurs when there is physical contact between prey and predator. In this case, the physical inactivity of the prey may prevent the continuation of the attack. The neural substrate of this response is not well known and the objective of the present study was to investigate the effect of cholinergic stimulation of hypothalamic regions on TI modulation in guinea pigs (Cavia porcellus). Microinjection of carbachol (1.0 microg/0.2 microL) into the anterior hypothalamus promoted an increase in the duration of TI episodes. Microinjection of carbachol into the ventro- and dorsomedial hypothalamus, however, promoted a reduced duration of TI episodes. Pretreatment with atropine (0.5 microg/0.2 microL) showed that the action of carbachol is mediated by muscarinic receptors in the anterior and ventromedial hypothalamus but not in the dorsomedial hypothalamus. The results suggest that the hypothalamic regions may play different roles in the organization of defensive behavioral responses such as TI.

Interaction between NO and oxytocin: influence on LHRH release.

Nitric oxide synthase (NOS)-containing neurons have been localized in various parts of the CNS. These neurons occur in the hypothalamus, mostly in the paraventricular and supraoptic nuclei and their axons project to the neural lobe of the pituitary gland. We have found that nitric oxide (NO) controls luteinizing hormone-releasing hormone (LHRH) release from the hypothalamus acting as a signal transducer in norepinephrine (NE)-induced LHRH release. LHRH not only releases LH from the pituitary but also induces sexual behavior. On the other hand, it is known that oxytocin also stimulates mating behavior and there is some evidence that oxytocin can increase NE release. Therefore, it occurred to us that oxytocin may also stimulate LHRH release via NE and NO. To test this hypothesis, we incubated medial basal hypothalamic (MBH) explants from adult male rats in vitro. Following a preincubation period of 30 min, MBH fragments were incubated in Krebs-Ringer bicarbonate buffer in the presence of various concentrations of oxytocin. Oxytocin released LHRH at concentrations ranging from 0.1 nM to 1 microM with a maximal stimulatory effect (P < 0.001) at 0.1 microM, but with no stimulatory effect at 10 microM. That these effects were mediated by NO was shown by the fact that incubation of the tissues with NG-monomethyl-L-arginine (NMMA), a competitive inhibitor of NOS, blocked the stimulatory effects. Furthermore, the release of LHRH by oxytocin was also blocked by prazocin, an alpha 1-adrenergic receptor antagonist, indicating that NE mediated this effect. Oxytocin at the same concentrations also increased the activity of NOS (P < 0.01) as measured by the conversion of [14C]arginine to citrulline, which is produced in equimolar amounts with NO by the action of NOS. The release of LHRH induced by oxytocin was also accompanied by a significant (P < 0.02) increase in the release of prostaglandin E2 (PGE2), a mediator of LHRH release that is released by NO. On the other hand, incubation of neural lobes with various concentrations of sodium nitroprusside (NP) (300 or 600 microM), a releaser of NO, revealed that NO acts to suppress (P < 0.01) the release of oxytocin. Therefore, our results indicate that oxytocin releases LHRH by stimulating NOS via NE, resulting in an increased release of NO, which increases PGE2 release that in turn induces LHRH release. Furthermore, the released NO can act back on oxytocinergic terminals to suppress the release of oxytocin in an ultrashort-loop negative feedback.

Interaction between NO and oxytocin: influence on LHRH release

Nitric oxide synthase (NOS)-containing neurons have been localized in various parts of the CNS. These neurons occur in the hypothalamus, mostly in the paraventricular and supraoptic nuclei and their axons project to the neural lobe of the pituitary gland. We have found that nitric oxide (NO) controls luteinizing hormone-releasing hormone (LHRH) release from the hypothalamus acting as a signal transducer in norepinephrine (NE)-induced LHRH release. LHRH not only releases LH from the pituitary but also induces sexual behavior.On the other hand, it is known that oxytocin also stimulates mating behavior and there is some evidence that oxytocin can increase NE release. Therefore, it occurred to us that oxytocin may also stimulate LHRH releave via NE and NO. To test this hypothesis, we incubated medial basal hypothalamic (MBH) explants from adult male rats in vitro. Following a preincubation period of 30 min, MBH fragments were incubated in Krebs-Ringer bicarbonate buffer in the presence of various concentrations of oxytocin. Oxytocin relesed LHRH at concentrations ranging from 0.1 nM to 1muM with a maximal stimulatory effect (P<0.001) at 0.1 muM, but with no stimulatory effect at 10 muM. That these effects were mediated by NO was shown by the fact that incubation of the tissues with NG-monomethyl-L-arginine (NMMA), a competitive inhibitor of NOS, blocked the stimulatory effects. Furthermore, the release of LHRH by oxytocin was also blocked by prazocin, an alpha1-adrenergic receptor antagonist, indicating that NE mediated this effect. Oxytocin at the same concentrations also increased the activity of NOS (P<0.01) as measured by the conversion of [14C]arginine to citrulline, which is produced in equimolar amounts with NO by the action of NOS. The release of LHRH induced by oxytocin was also accompanied by a significant (P<0.02) increase in the release of prostaglandin E2 (PGE2), a mediator of LHRH release that is released by NO. On the other hand, incubation of neural lobes with vaious concentrations of sodium nitroprusside (NP) (300 or 600 muM), a releaser of NO, revealed that NO acts to suppres (P<0.01) the release of oxytoxin. Therefore, our results indicate that oxytocin releases LHRH by stimulating NOS via NE, resulting in an increased release of NO, which increases PGE2 release that in turn induces LHRH release. Furthermore, the released NO can act back on oxytocinergic terminals to suppress the release of oxytocin in an ultrashort-loop negative feedback.

Oxytocin stimulates the release of luteinizing hormone-releasing hormone from medial basal hypothalamic explants by releasing nitric oxide.

Oxytocin induces mating behavior in rats of both sexes. Previous experiments revealed that progesterone-induced sex behavior in ovariectomized, estrogen-primed rats was caused by release of NO from NOergic neurons that stimulated the release of luteinizing hormone-releasing hormone (LHRH). The LHRH activated brain-stem neurons that initiated the lordosis reflex. We hypothesized that oxytocin might similarly release NO in the medial basal hypothalamic region that would stimulate release of LHRH into the hypophyseal portal vessels to release luteinizing hormone. To investigate this hypothesis, medial basal hypothalamic explants were preincubated in Krebs-Ringer bicarbonate buffer for 30 min, followed by a 30-min incubation in fresh Krebs-Ringer bicarbonate buffer containing the compounds to be tested. Oxytocin stimulated LHRH release 3- to 4-fold at the lowest concentration tested (10(-10) M). Values remained at a plateau as the concentration was increased to 10(-7) M and then declined in a concentration-dependent manner, so that there was no stimulation with a concentration of 10(-5) M. Oxytocin (10(-7) M) stimulated release of prostaglandin E2 into the medium, a finding consistent with a role of NO in the response. That NO indeed mediated the action of oxytocin was supported by blockade of the action of oxytocin by the competitive inhibitor of NO synthase (NOS), N(G)-monomethyl-L-arginine (300 microM). Furthermore, oxytocin (10(-9) to 10(-7) M) activated NOS as measured at the end of the experiments. Oxytocin appeared to act to stimulate norepinephrine terminals in the medial basal hypothalamus, which activated NOS by alpha1-adrenergic receptors, because prazocine, an alpha1 receptor blocker, inhibited the LHRH-releasing action of oxytocin. Finally, incubation of neural lobe explants with sodium nitroprusside, a NO releasor, revealed that nitroprusside (300-600 microM, but not 900 microM) inhibited oxytocin release. Therefore, the NO released by oxytocin also diffuses into the oxytocin neuronal endings and inhibits oxytocin release, forming a negative feedback loop. The results indicate that oxytocin is important not only in induction of mating, but also in stimulating LHRH release with subsequent luteinizing hormone discharge that plays a crucial role in reproduction.

Ibotenate lesion of the medial hypothalamus alters the salt intake and pressor responses to activation of the median preoptic nucleus in rats.

We investigated the influence of ibotenic acid lesions of the medial hypothalamus (MH) on salt appetite and arterial blood pressure responses induced by angiotensinergic and adrenergic stimulation of the median preoptic nucleus (MnPO) of rats. Previous injection of the adrenergic agonists norepinephrine, clonidine, phenylephrine, and isoproterenol into the MnPO of sham MH-lesioned rats caused no change in the sodium intake induced by ANG II. ANG II injected into the MnPO of MH-lesioned rats increased sodium intake compared with sham-lesioned rats. Previous injection of clonidine and isoproterenol increased, whereas phenylephrine abolished the salt intake induced by ANG II into the MnPO of MH-lesioned rats. Previous injection of norepinephrine and clonidine into the MnPO of sham MH-lesioned rats caused no change in the mean arterial pressure (MAP) induced by ANG II. Under the same conditions, previous injection of phenylephrine increased, whereas isoproterenol reversed the increase in MAP induced by angiotensin II (ANG II). ANG II injected into the MnPO of MH-lesioned rats induce a decrease in MAP compared with sham-lesioned rats. Previous injection of phenylephrine or norepinephrine into the MnPO of MH-lesioned rats induced a negative MAP, whereas pretreatment with clonidine or isoproterenol increased the MAP produced by ANG II injected into the MnPO of sham- or MH-lesioned rats. These data show that ibotenic acid lesion of the MH increases the sodium intake and pressor responses induced by the concomitant angiotensinergic, alpha 2 and beta adrenergic activation of the MnPO, whereas alpha 1 activation may have opposite effects. MH involvement in excitatory and inhibitory mechanisms related to sodium intake and MAP control is suggested.

Neurokinin A affects the tubero-hypophyseal gabaergic system.

We have studied the in vitro effects of neurokinin A (NKA) on anterior pituitary GABA concentration and GABA release from the mediobasal hypothalamus and the neurointermediate lobe of male and ovariectomized female (OVX) rats. NKA significantly decreased the anterior pituitary GABA concentration, while the presence of a specific anti-NKA serum in the incubation medium increased the GABA concentration in this gland. By contrast, NKA did not modify basal or K(+)-evoked GABA release from the mediobasal hypothalamus of male or OVX rats. However, NKA decreased basal and K(+)-evoked GABA release from the neurointermediate lobe. Since GABA inhibits both prolactin (PRL) secretion from the anterior pituitary and the release of several putative PRL-releasing factors from the neurointermediate lobe, the decrease in anterior pituitary GABA concentration and the reduction in tubero-hypophyseal GABAergic activity induced by NKA may contribute to the stimulatory effect of this peptide on PRL secretion.

Dipsogenic effect of pituitary adenylate cyclase activating polypeptide (PACAP38) injected into the lateral hypothalamus.

PACAP38 bilaterally injected in the vicinity of the perifornical lateral hypothalamus (pfLH) induced drinking behavior in rats. The animals (n = 12) drank 19.7 +/- 4.1 ml of water during the hour following PACAP38 microinjections (1 nmol/0.5 microliter). In the same rat sulpiride microinjections (45 nmol/0.5 microliter) had relatively mild effects (7.8 +/- 1.4 ml/h). The dipsogenic effects of sulpiride and PACAP38 were well correlated suggesting that both substances trigger drinking behavior activating the same hypothalamic mechanisms. Neither sulpiride nor PACAP38 promoted drinking when injected just 1.3 mm behind the effective zone. This negative result is an evidence of the neuroanatomical specificity of the dipsogenic effects of both substances. These preliminary results suggest that PACAP38 in the pfLH could be a neuropeptide regulating drinking behavior and perhaps body fluid volume and osmolarity and arterial blood pressure.

Effect of lateral hypothalamus lesions on the water and salt intake, and sodium and urine excretion induced by activation of the median preoptic nucleus in conscious rats.

In this study we investigated the influence of electrolytic lesion of the lateral hypothalamus (LH) on the water and salt appetite, and the natriuretic, diuretic and cardiovascular effects induced by angiotensinergic, cholinergic and noradrenergic stimulation of the median preoptic nucleus (MnPO) in rats. Male Holtzman rats were implanted with a cannula into the MnPO. Other groups of sham- and LH-lesioned rats received a stainless steel cannula implanted into the MnPO. ANGII injection into the MnPO induced water and sodium intake, and natriuretic, diuretic, pressor and tachycardic responses. Carbachol induced water intake, and natriuretic, pressor and bradycardic responses, whereas noradrenaline increased urine, sodium excretion and blood pressure, and induced bradycardia. In rats submitted to LH-lesion only, water and sodium intake was reduced compared with sham rats. LH lesion also reduced the sodium ingestion induced by ANGII (12 ng) into the MnPO. In LH-lesioned rats, the dipsogenic, diuretic and pressor responses induced by ANGII (12 ng), carbachol (2 nmol) and noradrenaline (20 nmol) injection into the MnPO were reduced. The same occurred with sodium excretion when carbachol (2 nmol) and noradrenaline (20 nmol) were injected into the MnPO of LH-lesioned rats, whereas ANGII (12 ng) induced an increase in sodium excretion. These data show that electrolytic lesion of the LH reduces fluid and sodium intake, and pressor responses to angiotensinergic, cholinergic and noradrenergic activation of the MnPO. LH involvement with MnPO excitatory and inhibitory mechanisms related to water and sodium intake, sodium excretion and cardiovascular control is suggested.

c-fos immunoreactivity in the brain following electrical or chemical stimulation of the medial hypothalamus of freely moving rats.

c-fos immunoreactivity was used to map brain areas in which neurons reacted either to electrical stimulation or to microinjection of the excitatory amino acid kainate and of the GABAA antagonist, SR-95531, applied to the medial hypothalamus of freely moving rats. All these stimulations induced flight behavior of moderate intensity. Immunoreactive cells were found within a radius of 0.5 mm around the stimulated area. Distally, clusters of labeled cells were found ipsilaterally in the piriform and entorhinal cortices, in several amygdaloid nuclei, in the bed nucleus of the stria terminalis, in the septo-hypothalamic nucleus, in the paraventricular, anterior and dorsomedial hypothalamic nuclei, the the paraventricular thalamic nucleus, in the dorsal periaqueductal gray extending to the cuneiform nucleus, and bilaterally in the supramammillary decussation and the locus coeruleus. The specificity of the brain areas thus labeled was indicated by the unilateral pattern of activation as well as by the different pattern obtained after control microinjection of saline. Therefore, these results are likely to provide sound information about the brain structures involved in defensive-aversive behavior evoked from the medial hypothalamus.

Regional distribution of Fos-like immunoreactivity in the rat brain after exposure to fear-inducing stimuli.

Fos protein immunohistochemistry was used to identify the neural substrate of fear/anxiety. The structures activated by exposure of Long Evans male rats (280-300 g) to the elevated plus-maze, a widely used animal model of anxiety, were compared with those activated by chemical stimulation of two aversive areas of the brain, the dorsal periaqueductal gray matter and the medial hypothalamus. Three different patterns of activation were obtained: Pattern 1 resulted from microinjection of the excitatory amino acid kainate (60 pmol; N = 5) or of the GABA(A) receptor antagonist SR-95531 (16 pmol; N = 3) into the dorsal periaqueductal gray matter and consisted mainly of caudal structures; Pattern 2 was observed after kainate injection (60 pmol; N = 4) into the medial hypothalamus and had a predominantly prosencephalic distribution; Pattern 3 extended from rostral to caudal brain regions and was induced by microinjection of either SR-95531 (16 pmol; N = 1) or kainate (120 pmol; N = 3) into the medial hypothalamus, as well as by 15-min exposure to the plus-maze (N = 3). Control animals were either injected with saline into the MH (N = 3) or the PAG (N = 3) or were exposed for 15 s to the elevated plus maze (N = 3) and exhibited no significant labeling. These results further support the participation of periventricular structures in the regulation of fear and aversion.

Regional distribution of Fos-like immunoreactivity in the rat brain after exposure to fear-inducing stimuli

Fos protein immunohistochemistry was used to identify the neural substrate of fear/anxiety. The structures activated by exposure of Long Evans male rats (280-300 g) to the elevated plus-maze, a widely used animal model of anxiety, were compared with those activated by chemical stimulation of two aversive areas of the brain, the dorsal periaqueductal gray matter and the medial hypothalamus. Three different patterns of activation were obtained: Pattern 1 resulted from microinjection of the excitatory amino acid kainate (60 pmol; N = 5) or of the GABA(A) receptor antagonist SR-95531 (16 pmol; N = 3) into the dorsal periaqueductal gray matter and consisted mainly of caudal structures; Pattern 2 was observed after kainate injection (60 pmol; N = 4) into the medial hypothalamus and had a predominantly prosencephalic distribution; Pattern 3 extended from rostral to caudal brain regions and was induced by microinjection of either SR-95531 (16 pmol; N = 1) or kainate (120 pmol; N = 3) into the medial hypothalamus, as well as by 15-min exposure to the plus-maze (N = 3). Control animals were either injected with saline into the MH (N = 3) or the PAG (N = 3) or were exposed for 15 s to the elevated plus maze (N = 3) and exhibited no significant labeling. These results further support the participation of periventricular structures in the regulation of fear and aversion.

Short-term changes in natripheric and hydrosmotic water fluxes across the skin and in urine production due to increases in the osmolarity of the external environment in the toad.

1. Sudden decreases in the osmotic gradient across the skin due to the replacement of water of the bath by 115 mM NaCl had no effect on water uptake of intact or hypophysectomized toads. 2. A concomitant decrease in the urine production was observed in intact but not in hypophysectomized animals. 3. Addition of amiloride chlorydrate (0.25 mM) to the 115 mM NaCl bath induced a significant decrease in water uptake both in intact and in hypophysectomized toads. 4. The osmotic permeability coefficient (LPD) increased significantly during the osmotic gradient reduction with 115 mM NaCl plus 0.25 mM amiloride or 230 mM sucrose in both groups. 5. No changes in the plasmatic osmolarity were detected during the development of these responses to the osmotic gradient reduction. 6. These results are consistent with the hypothesis of short-term changes in the natripheric and hydrosmotic fluxes of water across the skin and in urine production triggered by the osmotic gradient reduction. The possible participation of arginine vasotocin in these responses is discussed.

Effect of estrogen and progesterone priming on the uptake and release of serotonin and noradrenaline from the ventromedial hypothalamus.

Estradiol benzoate (EB) or EB plus progesterone (P) were administered to chronic ovariectomized rats in doses which are known to induce sexual receptivity. P was injected 48 hr after EB and the animals were decapitated 54 hr after E.B. The ventromedial hypothalamus (VMH) (excluding the arcuate-median eminence region) was dissected out and used to determine 3H-noradrenaline (3H-NA) and 3H-serotonin (3H-5-HT) uptake and release induced by high potassium concentration. Animals injected with oil were used as controls. VMH from EB (10 micrograms/kg) plus P (2 mg/kg) primed animals had higher release of 3H-NE than controls; uptake and release of 3H-5-HT was also augmented in these animals. EB (100 micrograms/kg) priming alone did not induce significant variations. The increase in 3H-NA release is interpreted as evidence of incremented activity in a 3H-NA system, facilitating sexual receptivity. The change in 3H-5-HT uptake and release is discussed in respect to the existence of an inhibitory serotonergic system modulating sexual behavior.