Dorsal periaqueductal gray stimulation facilitates anxiety-, but not panic-related, defensive responses in rats tested in the elevated T-maze

The escape response to electrical or chemical stimulation of the dorsal periaqueductal gray matter (DPAG) has been associated with panic attacks. In order to explore the validity of the DPAG stimulation model for the study of panic disorder, we determined if the aversive consequences of the electrical or chemical stimulation of this midbrain area can be detected subsequently in the elevated T-maze. This animal model, derived from the elevated plus-maze, permits the measurement in the same rat of a generalized anxiety- and a panic-related defensive response, i.e., inhibitory avoidance and escape, respectively. Facilitation of inhibitory avoidance, suggesting an anxiogenic effect, was detected in male Wistar rats (200-220 g) tested in the elevated T-maze 30 min after DPAG electrical stimulation (current generated by a sine-wave stimulator, frequency at 60 Hz) or after local microinjection of the GABA A receptor antagonist bicuculline (5 pmol). Previous electrical (5, 15, 30 min, or 24 h before testing) or chemical stimulation of this midbrain area did not affect escape performance in the elevated T-maze or locomotion in an open-field. No change in the two behavioral tasks measured by the elevated T-maze was observed after repetitive (3 trials) electrical stimulation of the DPAG. The results indicate that activation of the DPAG caused a short-lived, but selective, increase in defensive behaviors associated with generalized anxiety.

Behavioral meaningful opioidergic stimulation activates kappa receptor gene expression

The periaqueductal gray (PAG) has been reported to be a location for opioid regulation of pain and a potential site for behavioral selection in females. Opioid-mediated behavioral and physiological responses differ according to the activity of opioid receptor subtypes. The present study investigated the effects of the peripheral injection of the kappa-opioid receptor agonist U69593 into the dorsal subcutaneous region of animals on maternal behavior and on Oprk1 gene activity in the PAG of female rats. Female Wistar rats weighing 200-250 g at the beginning of the study were randomly divided into 2 groups for maternal behavior and gene expression experiments. On day 5, pups were removed at 7:00 am and placed in another home cage that was distant from their mother. Thirty minutes after removing the pups, the dams were treated with U69593 (0.15 mg/kg, sc) or 0.9% saline (up to 1 mL/kg) and after 30 min were evaluated in the maternal behavior test. Latencies in seconds for pup retrieval, grouping, crouching, and full maternal behavior were scored. The results showed that U69593 administration inhibited maternal behavior (P < 0.05) because a lower percentage of kappa group dams showed retrieval of first pup, retrieving all pups, grouping, crouching and displaying full maternal behavior compared to the saline group. Opioid gene expression was evaluated using real-time reverse-transcription polymerase chain reaction (RT-PCR). A single injection of U69593 increased Oprk1 PAG expression in both virgin (P < 0.05) and lactating female rats (P < 0.01), with no significant effect on Oprm1 or Oprd1 gene activity. Thus, the expression of kappa-opioid receptors in the PAG may be modulated by single opioid receptor stimulation and behavioral meaningful opioidergic transmission in the adult female might occur simultaneously to specific changes in gene expression of kappa-opioid receptor subtype. This is yet another alert for the complex role of the opioid ...

Involvement of midbrain tectum neurokinin-mediated mechanisms in fear and anxiety

Electrical stimulation of midbrain tectum structures, particularly the dorsal periaqueductal gray (dPAG) and inferior colliculus (IC), produces defensive responses, such as freezing and escape behavior. Freezing also ensues after termination of dPAG stimulation (post-stimulation freezing). These defensive reaction responses are critically mediated by γ-aminobutyric acid and 5-hydroxytryptamine mechanisms in the midbrain tectum. Neurokinins (NKs) also play a role in the mediation of dPAG stimulation-evoked fear, but how NK receptors are involved in the global processing and expression of fear at the level of the midbrain tectum is yet unclear. The present study investigated the role of NK-1 receptors in unconditioned defensive behavior induced by electrical stimulation of the dPAG and IC of male Wistar rats. Spantide (100 pmol/0.2 μL), a selective NK-1 antagonist, injected into these midbrain structures had anti-aversive effects on defensive responses and distress ultrasonic vocalizations induced by stimulation of the dPAG but not of the IC. Moreover, intra-dPAG injections of spantide did not influence post-stimulation freezing or alter exploratory behavior in rats subjected to the elevated plus maze. These results suggest that NK-1 receptors are mainly involved in the mediation of defensive behavior organized in the dPAG. Dorsal periaqueductal gray-evoked post-stimulation freezing was not affected by intra-dPAG injections of spantide, suggesting that NK-1-mediated mechanisms are only involved in the output mechanisms of defensive behavior and not involved in the processing of ascending aversive information from the dPAG.

Contrasting effects of nitric oxide and corticotropin- releasing factor within the dorsal periaqueductal gray on defensive behavior and nociception in mice

The anxiogenic and antinociceptive effects produced by glutamate N-methyl-D-aspartate receptor activation within the dorsal periaqueductal gray (dPAG) matter have been related to nitric oxide (NO) production, since injection of NO synthase (NOS) inhibitors reverses these effects. dPAG corticotropin-releasing factor receptor (CRFr) activation also induces anxiety-like behavior and antinociception, which, in turn, are selectively blocked by local infusion of the CRF type 1 receptor (CRFr1) antagonist, NBI 27914 [5-chloro-4-(N-(cyclopropyl)methyl-N-propylamino)-2-methyl-6-(2,4,6-trichlorophenyl)aminopyridine]. Here, we determined whether i) the blockade of the dPAG by CRFr1 attenuates the anxiogenic/antinociceptive effects induced by local infusion of the NO donor, NOC-9 [6-(2-hydroxy-1-methyl-2-nitrosohydrazino)-N-methyl-1-hexanamine], and ii) the anxiogenic/antinociceptive effects induced by intra-dPAG CRF are prevented by local infusion of Nω-propyl-L-arginine (NPLA), a neuronal NOS inhibitor, in mice. Male Swiss mice (12 weeks old, 25-35 g, N = 8-14/group) were stereotaxically implanted with a 7-mm cannula aimed at the dPAG. Intra-dPAG NOC-9 (75 nmol) produced defensive-like behavior (jumping and running) and antinociception (assessed by the formalin test). Both effects were reversed by prior local infusion of NBI 27914 (2 nmol). Conversely, intra-dPAG NPLA (0.4 nmol) did not modify the anxiogenic/antinociceptive effects of CRF (150 pmol). These results suggest that CRFr1 plays an important role in the defensive behavior and antinociception produced by NO within the dPAG. In contrast, the anxiogenic and antinociceptive effects produced by intra-dPAG CRF are not related to NO synthesis in this limbic midbrain structure.

Morphine infusions into the rostrolateral periaqueductal gray affect maternal behaviors

It is well established that morphine inhibits maternal behaviors. Previous studies by our group have shown activation of the rostrolateral periaqueductal gray (rlPAG) upon inhibition-intended subcutaneous injections of morphine. In this context, we demonstrated that a single naloxone infusion into the rlPAG, following this opioid-induced inhibition, reactivated maternal behaviors. Since these data were obtained by using peripheral morphine injections, the present study was designed to test whether morphine injected directly into the rlPAG would affect maternal behaviors. Our hypothesis that morphine acting through the rlPAG would disrupt maternal behaviors was confirmed with a local infusion of morphine. The mothers showed shorter latency for locomotor behavior to explore the home cage (P = 0.049). Inhibition was especially evident regarding retrieving (P = 0.002), nest building (P = 0.05) and full maternal behavior (P = 0.023). These results support the view that opioidergic transmission plays a behaviorally meaningful inhibitory role in the rostrolateral PAG.

The synergetic modulation of the excitability of central gray matter by a neuropeptide: two protocols using excitation waves in chick retina

The isolated chick retina provides an in vitro tissue model, in which two protocols were developed to verify the efficacy of a peptide in the excitability control of the central gray matter. In the first, extra-cellular potassium homeostasis is challenged at long intervals and in the second, a wave is trapped in a ring of tissue causing the system to be under self-sustained challenge. Within the neuropil, the extra-cellular potassium transient observed in the first protocol was affected from the initial rising phase to the final concentration at the end of the five-minute pulse. There was no change in the concomitants of excitation waves elicited by the extra-cellular rise of potassium. However, there was an increase on the elicited waves latency and/or a rise in the threshold potassium concentration for these waves to appear. In the second protocol, the wave concomitants and the propagation velocity were affected by the peptide. The results suggest a synergetic action of the peptide on glial and synaptic membranes: by accelerating the glial Na/KATPase and changing the kinetics of the glial potassium channels, with glia tending to accumulate KCl. At the same time, there is an increase in potassium currents through nerve terminals.

Retinas de pinto isoladas proporcionam um modelo de tecidos in vitro, para o qual dois protocolos foram desenvolvidos para verificar a eficácia de um peptídeo no controle da excitabilidade da matéria cinzenta central. No primeiro, a homeostase do potássio extra-celular é desafiada por intervalos longos (1 hora) e no segundo, uma onda é capturada em um anel de tecido, de tal maneira que o sistema permaneça em estado de desafio auto-sustentado. Dentro da neuropil, o transiente de potássio extra-celular observado no primeiro protocolo foi afetado da fase de início de aumento à concentração final, ao final do pulso de cinco minutos. Não há mudanças nos parâmetros concomitantes das ondas de excitação geradas pelo aumento do potássio extra-celular. Entretanto, houve um aumento da latência das ondas geradas e/ou um aumento no nível de concentração de potássio necessário para gerar a onda. No segundo protocolo, os parâmetros concomitantes da onda e sua velocidade de propagação foram afetados pelo peptídeo. Os resultados sugerem uma ação sinergética do peptídeo nas membranas gliais e sinápticas: acelerando o Na/KATPase glial e mudando a cinética dos canais de potássio gliais, com a glia tendendo a acumular KCl. Nesse período, não há aumento nas correntes de potássio nas terminações nervosas.

Maternal aggression in Wistar rats: effect of 5-HT2A/2C receptor agonist and antagonist microinjected into the dorsal periaqueductal gray matter and medial septum

The objective of the present study was to assess the role of the 5-HT2A/2C receptor at two specific brain sites, i.e., the dorsal periaqueductal gray matter (DPAG) and the medial septal (MS) area, in maternal aggressive behavior after the microinjection of either a 5-HT2A/2C receptor agonist or antagonist. Female Wistar rats were microinjected on the 7th postpartum day with the selective agonist alpha-methyl-5-hydroxytryptamine maleate (5-HT2A/2C) or the antagonist 5-HT2A/2C, ketanserin. The agonist was injected into the DPAG at 0.2 (N = 9), 0.5 (N = 10), and 1.0 æg/0.2 æl (N = 9), and the antagonist was injected at 1.0 æg/0.2 æl (N = 9). The agonist was injected into the medial septal area (MS) at 0.2 (N = 9), 0.5 (N = 7), and 1.0 æg/0.2 æl (N = 6) and the antagonist was injected at 1.0 æg/0.2 æl (N = 5). For the control, saline was injected into the DPAG (N = 7) and the MS (N = 12). Both areas are related to aggressive behavior and contain a high density of 5-HT receptors. Non-aggressive behaviors such as horizontal locomotion (walking) and social investigation and aggressive behaviors such as lateral threat (aggressive posture), attacks (frontal and lateral), and biting the intruder were analyzed when a male intruder was placed into the female resident's cage. For each brain area studied, the frequency of the behaviors was compared among the various treatments by analysis of variance. The results showed a decrease in maternal aggressive behavior (number of bites directed at the intruder) after microinjection of the agonist at 0.2 and 1.0 æg/0.2 æl (1.6 ± 0.7 and 0.9 ± 0.3) into the DPAG compared to the saline group (5.5 ± 1.1). There was no dose-response relationship with the agonist. The present findings suggest that the 5-HT2A/2C receptor agonist has an inhibitory effect on maternal aggressive behavior when microinjected into the DPAG and no effect when microinjected into the MS. Ketanserin (1.0 æg/0.2 æl) decreased locomotion when microinjected into the DPAG and MS, but did not affect aggressive behavior. We interpret these findings as evidence for a specific role of 5-HT2A/2C receptors in the DPAG in the inhibition of female aggressive behavior, dissociated from those on motor activity.

The brain decade in debate: II. Panic or anxiety? From animal models to a neurobiological basis

This article is a transcription of an electronic symposium sponsored by the Brazilian Society of Neuroscience and Behavior (SBNeC). Invited researchers from the European Union, North America and Brazil discussed two issues on anxiety, namely whether panic is a very intense anxiety or something else, and what aspects of clinical anxiety are reproduced by animal models. Concerning the first issue, most participants agreed that generalized anxiety and panic disorder are different on the basis of clinical manifestations, drug response and animal models. Also, underlying brain structures, neurotransmitter modulation and hormonal changes seem to involve important differences. It is also common knowledge that existing animal models generate different types of fear/anxiety. A challenge for future research is to establish a good correlation between animal models and nosological classification

Effect of microinjections of 5-hydroxytryptamine and adrenaline in central grey on pain responsiveness during acute food deprivation in conscious rats.

To study the effects of microinjections of 5 hydroxytryptamine and adrenaline in central grey on pain responsiveness during acute food deprivation, experiments were conducted in nine male rats. Microinjections of 5 HT (10 micrograms/microliter) and adrenaline (10 micrograms/microliter) were given in central grey before and at the end of 6, 12, 18 and 24 hr food deprivation and the effects on pain threshold, cardiorespiratory parameters and body temperature were noted. Observations showed that 5 HT increased the pain threshold (antinociception) significantly (P < 0.05) with no change in cardiorespiratory response and body temperature, adrenaline did not alter pain threshold with no change in cardiorespiratory response and body temperature. The observations suggest the possible existence of two types of monoaminergic receptors or pathways in the central grey.

Behavioral effects of "vehicle" microinjected into the dorsal periaqueductal grey of rats tested in the elevated plus maze

To investigate the behavioral effects of different vehicles microinjected into the dorsal periaqueductal grey (DPAG) of male Wistar rats, weighing 200-250 g, tested in the elevated plus maze, animals were implanted with cannulas aimed at this structure. One week after surgery the animals received microinjections into the DPAG of 0.9 per cent (w/v) saline, 10 per cent (v/v) dimethyl sulfoxide (DMSO), 2 per cent (v/v) Tween-80, 10 per cent (v/v) propylene glycol, or synthetic cerebrospinal fluid (CSF). Ten min after the injection (0.5 mul) the animals (N = 8-13/group) were submitted to the elevated plus maze test. DMSO significantly increased the number of entries into both the open and enclosed arms when compared to 0.9 per cent saline (2.7 ñ 0.8 and 8.7 ñ 1.3 vs 0.8 ñ 0.3 and 5.1 ñ 0.9, respectively, Duncan test, P<0.05), and tended to increase enclosed arm entries as compared to 2 per cent Tween-80 (8.7 ñ 1.3 vs 5.7 ñ 0.9, Duncan test, P

Participation of the periaqueductal gray matter in the modulation of tonic immobility in the guinea pig

Unilateral microinjection of carbachol (CCh, 1 microgram/0.2 microliter) into the specific sites in the ventral and ventrolateral portions of the periaqueductal gray (PAG) matter, which is known to be involved in analgesia, increases the duration of restraint-induced tonic immobility (TI) episodes induced in 23 adult male guinea pigs (Cavia porcellus). Mean duration of TI episodes was 107 +/- 16.38 s in the control group and increased to 220.7 +/- 40.24 s in the group microinjected with CCh. The potentiating effect of carbachol on TI duration was blocked by pretreatment with atropine (7.6 micrograms/0.4 microliter). These data suggest that PAG and the cholinergic system are involved in the modulation of the motor inhibition characteristic of TI which may be activated by the same stimuli that induce defensive analgesia.

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.

Midbrain adrenergic mechanisms modulating flight behaviour induced by hypothalamic stimulation.

The present study was carried out in ten cats of either sex. Flight response was obtained by electrical stimulation of dorsomedial regions of preoptic area (A13-14.5, L3.5 V-3.5 to -3.7) and lateral hypothalamic regions (A12.5, L2.5-3.5, V-3.7). It consisted of a goal directed attempt to get out of the cage with a vigorous leaping to foot. Norepinephrine when microinjected in 10 micrograms doses into pretectal area of midbrain (A3.5, 3.0, V+1.0 to +1.5 mm) significantly lowered the mean current strength from 640uA to 420uA; clonidine, an alpha-2 agonist in 5 micrograms dose when microinjected into the same locus also significantly lowered the mean current strength to the same level. On the other hand yohimbine, an alpha-2 blocker in 5 micrograms dose when microinjected in to the same locus significantly increased the mean current strength from 640 to 970 uA. These results indicate that hypothalamically induced flight response is mediated via the alpha-2 adrenoceptive mechanism operating at the midbrain level. Control microinjection of normal saline and propylene glycol in similar volumes failed to produce any changes in current strength.

Strain-dependent effects of morphine injected into the periaqueductal gray area of mice

Microinjection of morphine (0.31,1.25 and 5.0 *g) into the periaqueductal gray area (PAG) of C57BL/6 (C57) and DBA/2 (DBA) mice increased the pain threshold in the tail-flick test. The highest dose also caused a behavioral reaction in both strains characterized by periods of immobility laternating with explosive motor behavior. In the DBA strain, the analgesic effect was demonstrated with all doses of morphine,with in the C57 strain only the highest dose induced analgesia. DBA mice presented a decrease in activity with the lowest dose of morphine, whereas in the C57 strain, this effect was obtained only with the highest dose of morphine. These data corroborate at the PAG level the results of other studies which have shown that central and peripheral injections of morphine procedure analgesia and alter motor activity in C57 and DBA strains. They also confirm that these two strains of mice present genotype-dependent differences in sensitivity to opioids as determined after injections of morphine into the PAG

Anxiolytic effect of midazolam microinjected into the dorsal periaqueductal grey area of rats

In order to investigate the role of the dorsal periaqueductal grey (DPAG) area in the anxiolytic effect of benzodiazepines male Wistar rats (N=10), weighing 200-250 g at the time of surgery, were microinjected into this structure with midazolam (80 nmol) and submitted to the elevated plus-maze, an ethologically based model of anxiety. Midazolam significantly increased the percentage of open arm entries from 32.4 ñ 4.6 (control) to 49.5 ñ 3.0 and of time spent in the open arms from 21.0 ñ 4.5 (control) to 35.6 ñ4.8 without affecting the total number of entries into either open or enclosed arms. This effect typifies an anxiolytic effect in the test and was antagonized by the benzodiazepine receptor antagonist flumazenil (80 nmol) microinjection. Microinjection of flumazenil alone had no effect. These results provide additional evidence for the participation of the DPAG in the physiopathology of anxiety and suggest that it that it may be a site for the anxiolytic effect of systemically injected benzodiazepines

Anxiolytic effect of kynuremic acid microinjected into the dorsal periaqueductal gray matter of rats placed in the elevated plus-mase test

The effect of kynurenic acid (20 to 19=60 nmol) microinkected into the dorsal periaqueductal gray matter was measured in rats placed in a elevated plus-maze. Microinjection of 160 nmol of kynurenic acid increased the percentages of open arm entries and of time spent in the open arms. Both of these measures may be considered indexes of anciolysis. Although kynurenic acid also invreased the total number of entries, analysis of covariance shows that the increase in open arm entries is independent of the effect on closed arm entries. Thus, the anxiolytic effect of kynurenic acid detected in the elevated plus-maze strengthens the proposal that glutamatergic neurons of the dorsal periaqueductal gray matter paly an important role in anxiety

Microinjection of D-2-amino-7-phosphonoheptanoate into the dorsal periaqueductal gray matter reduces the pressor response to, glutamate injected at the same site

The current study assesses the influence of the N-methil-D-aspartate (NMDA) receptor antagonist D-2-amino-7-phosphonohepatanoate (AP7) on the pressor effect of glutamate microinjected into the dorsal periaqueductal gray matter (DPAG) of urethane-anesthetized rats. Glutamate (20, 40 and 80 nmol/site) caused dose-related reproducible increases in systolic and diastolic blood pressure. Microinjection of saline into the DPAG did not alter the pressor effects of glutamate (80 nmol/site). Similar pretreatment with AP7 (2nmol/site) significantly (P < 0.05) attenuated the pressor effects of glutamate from ñ26.5 ñ 7.0 to +3.4 ñ 3.3 mmHg (sistolic blood pressure). We conclude that the pressor efffect of glutamate in the DPAG is mediated largely by activation of NMDA receptors

Demência na hemodiálise: uso do WAIS e do MMPI / Hemodialysis dementia: WAIS and MMPI use

No SENUR do Hospital das Clínicas da U.F.M.G., o conteúdo de alumínio da água de torneira usada para compôr o dializado, revelou níveis acima do valor permitido. Quatorze pacientes com insuficiência renal crônica que se submetiam a hemodilise, em média 3 vezes por semana, foram estudados através do teste de inteligência para adultos Wechsler Bellevue (WAIS) e do Inventário Multifásico Minesota de Personalidade - (MMPI). Encontrou-se indicadores de deficiências cognitivas e emocionais próprias de encefalopatia da diálise, em 4 pacientes. O WAIS mostrou-se mais sensível para detectar este quadro clínico, principalmente na parte de execuçäo nos subtestes de Repetiçäo de Números, Número-Símbolo, Cubos, Composiçäo de Objetos e Arranjo de Figuras. Supoem-se uma correlaçäo entre a deposiçäo na substância cinzenta do cérebro do alumínio da água, ingerido através de medicamentos e a encefalopatía da diálise