F.G. Graeff

GABA mediation of the anti-aversive action of minor tranquilizers

Earlier observations have shown that systematically injected minor tranquilizers decrease the aversive consequences of electrical stimulation of the dorsal periaqueductal gray (DPAG) matter of the rat brain. In order to verify if these drugs can act directly on the DPAG, chlordiazepoxide (CDP) and pentobarbital (PB) were locally injected into the dorsal midbrain of rats chronically implanted with chemitrodes, allowing electrical stimulation of the same brain area. Microinjection of doses of 0.16 and 0.32 mumol of CDP and 0.16 mumol of PB significantly increased the threshold electrical current including flight behavior by stimulating the dorsal midbrain. Flight behavior was measured by the number of times rats crossed dividing line while running from one compartment of a shuttle-box to the other. The same effect was caused by the intracerebral injection of 0.32 and 0.64 mumol of the inhibitory neurotransmitter, gamma-aminobutyric acid (GABA). Conversely, local injection of the GABA antagonists. bicuculline (5-20) nmol) or picrotoxin (0.3 and 0.6 nmol), into the dorsal midbrain induced flight behavior, like the electrical stimulation. On the other hand, the glycine antagonist, strychnine (40 nmol) caused convulsive behavior only, while the intracerebral injection of the cholinergic agonist, carbachol (10-40 nmol), increased locomotion, sniffing and turning behavior, but did not induce flight. Pretreatment with locally injected GABA (0.64 mumol) antagonized the aversive effect of either bicuculline (10 nmol) or picrotoxin (0.3 nmol), whereas CDP (0.32 mumol) antagonised bicuculline only and PB (0.16 mumol) was ineffective against either bicuculline or picrotoxin. These results suggest that minor tranquilizers act directly upon the DPAG by enhancing the tonic inhibitory influence of endogenous GABA. This action may underly the antiaversive affects of these drugs.

Anxiolytic effect in the elevated plus-maze of the NMDA receptor antagonist AP7 microinjected into the dorsal periaqueductal grey

In order to localise the often reported anxiolytic action of N-methyl-D-aspartate (NMDA) receptor antagonists, 2-amino-7-phosphonoheptanoic acid (AP7) was injected into the dorsal periaqueductal grey (DPAG) of rats exposed to the elevated plus-maze model of anxiety. Doses of 0.2, 2 and 20 nmol AP7 caused a dose-dependent increase in the percentage of open arm entries, the effect of the last two doses being significantly different from control. A non-significant tendency to increase the percentage of time spent on the open arms of the maze was also noticed. In contrast, the total number of entries into either the open or enclosed arms was not affected. Injections of AP7 localized outside the DPAG were ineffective. Therefore, microinjection of AP7 into the DPAG caused a selective anxiolytic effect in the elevated plusmaze. It may be suggested that the DPAG is a site of the anxiolytic action of NMDA antagonists reported following systemic administration.

Anti-aversive role of serotonin in the dorsal periaqueductal grey matter

Microinjection of 5, 10, and 20 nmol serotonin (5-HT) and of 0.5, 1, and 2 nmol 5-methoxy-N, N-dimethyltryptamine (5-MeODMT) into the dorsal midbrain of rats bearing chronically implanted chemitrodes raised the electrical threshold for inducing escape behaviour following stimulation of the dorsal periaqueductal grey matter (DPAG). Linear regressions of log dose against drug-induced increase in aversive threshold were obtained for 5-HT and 5-MeODMT. The 5-MeODMT dose-effect curve was steeper and lay to the left of the 5-HT dose-effect curve. Local pre-treatment with 10 nmol metergoline or ketanserin blocked the anti-aversive effect of 10 nmol 5-HT, whereas pre-treatment with 100 nmol zimelidine potentiated this effect of 5-HT. The same dose of zimelidine raised the aversive threshold when given alone. These results suggest that 5-HT plays an inhibitory role in the DPAG controlling aversion, probably mediated by 5-HT2 receptors.

Effect of chlorimipramine and maprotiline on experimental anxiety in humans

In order to assess the role played by serotonin (5-HT) and noradrenaline in anxiety, four groups of healthy volunteers were given 25 mg of the selective inhibitor of 5-HT uptake chlorimipramine, 50 mg of the selective inhibitor of noradrenaline uptake maprotiline, 1 mg of the benzodiazepine anxiolytic lorazepam or placebo, and submitted to a simulated public speaking (SPS) test, consisting of speaking in front of a videocamera. Subjective anxiety was evaluated by the visual analog mood scale (VAMS) of Norris as well as by the state-trait anxiety inventory (STAI) of Spielberger. Chlorimipramine enhanced SPS-induced anxiety, whereas maprotiline and lorazepam reduced anxiety during as well as outside the test period. Mental and physical sedation (VAMS) were increased by either maprotiline or lorazepam. In a scale of bodily symptoms, chlorimipramine tended to increase muscle tension, agitation and palpitation, whereas maprotiline caused lethargy. The rise in blood pressure induced by the SPS procedure outlasted the period of stress in the group treated with chlorimipramine. In contrast, the SPS-induced increase in heart rate was enhanced by lorazepam. Chlor imipramine and maprotiline reduced salivation to the same extent. Pupillary diameter, however, was significantly increased by chlorimipramine alone. It may be tentatively sug gested that the proanxiogenic effect of chlorimipramine is related to changes in central 5-HT neurotransmission while the anxiolytic effect of maprotiline is associated with alteration of noradrenergic mechanisms. Increased peripheral sympathetic tone may also contribute to the proanxiety action of chlorimipramine.

Early life protein malnutrition changes exploration of the elevated plus-maze and reactivity to anxiolytics.

In order to investigate whether protein malnutrition in early life causes lasting changes in reactivity to anxiolytic drugs, exploration of the elevated plus-maze was used. Rat dams during lactation (21 days) and pups after weaning until day 49 of life were fed on 8% casein diet (M rats), while their well-nourished controls received 25% casein (W rats). From day 50 on all animals ate the same balanced diet. Experiments started on day 70. Under the non-drug condition, M rats tended to explore the open arms of the maze relatively more than W rats. Diazepam (0.5–5 mg/kg, IP) dose-dependently increased the percentage of open/total arm entries without significantly affecting the total number of arm entries in W rats. This selective anxiolytic effect of diazepam was considerably smaller in M rats. Ipsapirone (0.5–5 mg/kg) caused a similar though less pronounced anxiolytic effect in W rats, whereas the drug decreased both the % open/total and total arm entries in M rats. In contrast, ritanserin (0.05–1 mg/kg) significantly increased the % open/total arm entries in M rats only, though not in a dose-dependent way. Isamoltane (2.5–20 mg/kg) was ineffective on both M and W rats. These results indicate that early protein malnutrition causes long-lasting alterations in brain systems regulating emotional behaviour.

Role of benzodiazepine receptors located in the dorsal periaqueductal grey of rats in anxiety

Studies with electrical brain stimulation suggest that the dorsal periaqueductal grey matter (DPAG) is related to anxiety and to the anti-aversive effects of benzodiazepines (BZD) compounds. However, direct stimulation of the brain may prevent conclusions about the role of specific regions in the control of normal behaviour. In the present study we employed the elevated plus-maze, an ethologically based model of anxiety, to investigate the role of BZD receptors located in the DPAG in anxiety and in the anxiolytic effect of systemically injected BZD. The results showed that midazolam (20–80 nmol), a BZD agonist, dose-dependently increased the percentage of entries and time spent in open arms when microinjected into the DPAG. The effect of midazolam (80 nmol) was antagonized by flumazenil (80 nmol), a BZD antagonist, microinjected into the DPAG 10 min before the agonist. FG 7142 (20–80 nmol), a BZD partial inverse agonist, decreased time spent in open arms at the dose of 40 nmol and the number of open arms entries at all doses when microinjected into the DPAG. The microinjection of flumazenil (80 nmol) into the DPAG failed to antagonize the anxiolytic effect of systemically injected diazepam (2.5 mg/kg). These results strengthen the idea of an involvement of BZD receptors located in the DPAG with anxiety. They also suggest that the DPAG is not the only structure responsible for the anxiolytic effects of systemically injected BZD.

Pharmacology of human experimental anxiety

This review covers the effect of drugs affecting anxiety using four psychological procedures for inducing experimental anxiety applied to healthy volunteers and patients with anxiety disorders. The first is aversive conditioning of the skin conductance responses to tones. The second is simulated public speaking, which consists of speaking in front of a video camera, with anxiety being measured with psychometric scales. The third is the Stroop Color-Word test, in which words naming colors are painted in the same or in a different shade, the incongruence generating a cognitive conflict. The last test is a human version of a thoroughly studied animal model of anxiety, fear-potentiated startle, in which the eye-blink reflex to a loud noise is recorded. The evidence reviewed led to the conclusion that the aversive conditioning and potentiated startle tests are based on classical conditioning of anticipatory anxiety. Their sensitivity to benzodiazepine anxiolytics suggests that these models generate an emotional state related to generalized anxiety disorder. On the other hand, the increase in anxiety determined by simulated public speaking is resistant to benzodiazepines and sensitive to drugs affecting serotonergic neurotransmission. This pharmacological profile, together with epidemiological evidence indicating its widespread prevalence, suggests that the emotional state generated by public speaking represents a species-specific response that may be related to social phobia and panic disorder. Because of scant pharmacological data, the status of the Stroop Color-Word test remains uncertain. In spite of ethical and economic constraints, human experimental anxiety constitutes a valuable tool for the study of the pathophysiology of anxiety disorders.

Decreased left temporal lobe volume of panic patients measured by magnetic resonance imaging

Reported neuroimaging studies have shown functional and morphological changes of temporal lobe structures in panic patients, but only one used a volumetric method. The aim of the present study was to determine the volume of temporal lobe structures in patients with panic disorder, measured by magnetic resonance imaging. Eleven panic patients and eleven controls matched for age, sex, handedness, socioeconomic status and years of education participated in the study. The mean volume of the left temporal lobe of panic patients was 9% smaller than that of controls (t21 = 2.37, P = 0.028). In addition, there was a trend (P values between 0.05 and 0.10) to smaller volumes of the right temporal lobe (7%, t21 = 1.99, P = 0.06), right amygdala (8%, t21 = 1.83, P = 0.08), left amygdala (5%, t21 = 1.78, P = 0.09) and left hippocampus (9%, t21 = 1.93, P = 0.07) in panic patients compared to controls. There was a positive correlation between left hippocampal volume and duration of panic disorder (r = 0.67, P = 0.025), with recent cases showing more reduction than older cases. The present results show that panic patients have a decreased volume of the left temporal lobe and indicate the presence of volumetric abnormalities of temporal lobe structures.

Anxiolytic effect of glycine antagonists microinjected into the dorsal periaqueductal grey

To investigate if blockade of the modulatory glycine site of NMDA receptors in the dorsal periaqueductal grey (DPAG) would produce anxiolytic effects, groups of 9–14 rats received microinjections into this structure of 7-chloro-kynurenic acid (7-Cl-KY, 4 and 8 nmol) or 3-amino-1-hydroxypyrrolid-2-one (HA-966, 30 or 100 nmol), two selective antagonists at the strychnine-insensitive glycine modulatory site, and were submitted to the elevated plus-maze, an ethologically based animal model of anxiety. Both drugs increased the percentage of entries and of time spent in open arms as compared to rats receiving isotonic saline. Injections of the active compounds outside the DPAG were not effective. In another experiment microinjections of 7-Cl-KY (8 nmol) and HA-966 (100 nmol) into the DPAG raised the threshold of aversive electrical stimulation of the rat DPAG. These results indicate that microinjections of 7-Cl-KY and HA-966 into the DPAG cause anxiolytic effects in two different models of anxiety and support the proposal that NMDA-mediated neurotransmission in the DPAG may be related to anxiety and panic.

Ansiedade, pânico e o eixo hipotálamo-pituitária-adrenal

OBJECTIVE: This article focuses on the differential activation of the hypothalamic-pituitary-adrenal axis in generalized anxiety disorder and panic disorder. METHOD: The results of recently reported reviews of the literature are summarized and discussed. RESULTS: The results of experimental studies that assayed adrenocorticotropic hormone, cortisol and prolactin show that real-life panic attacks, as well as those induced by selective panicogenic agents such as lactate and carbon dioxide, do not activate the hypothalamic-pituitary-adrenal axis. Agonists of the cholecystokinin receptor B such as the cholecystokinin-4 peptide and pentagastrin increase stress hormones regardless of the occurrence of a panic attack and, thus, seem to activate the hypothalamic-pituitary-adrenal axis directly. The benzodiazepine antagonist flumazenil does not increase stress hormones, but this agent does not reliably induce panic attacks. Pharmacological agents that increase anxiety in both normal people and panic patients (caffeine, yohimbine, serotonergic agonists) raise stress hormone levels. CONCLUSIONS: In addition to the differences in symptomatology and pharmacological response, generalized anxiety disorder and panic disorder affect stress hormones in distinct ways. While anticipatory anxiety and generalized anxiety disorder activate both the hypothalamic-pituitary-adrenal and the sympathoadrenal axes, panic attack causes major sympathetic activation, but has little effect on the hypothalamic-pituitary-adrenal axis.OBJECTIVE This article focuses on the differential activation of the hypothalamic-pituitary-adrenal axis in generalized anxiety disorder and panic disorder. METHOD The results of recently reported reviews of the literature are summarized and discussed. RESULTS The results of experimental studies that assayed adrenocorticotropic hormone, cortisol and prolactin show that real-life panic attacks, as well as those induced by selective panicogenic agents such as lactate and carbon dioxide, do not activate the hypothalamic-pituitary-adrenal axis. Agonists of the cholecystokinin receptor B such as the cholecystokinin-4 peptide and pentagastrin increase stress hormones regardless of the occurrence of a panic attack and, thus, seem to activate the hypothalamic-pituitary-adrenal axis directly. The benzodiazepine antagonist flumazenil does not increase stress hormones, but this agent does not reliably induce panic attacks. Pharmacological agents that increase anxiety in both normal people and panic patients (caffeine, yohimbine, serotonergic agonists) raise stress hormone levels. CONCLUSIONS In addition to the differences in symptomatology and pharmacological response, generalized anxiety disorder and panic disorder affect stress hormones in distinct ways. While anticipatory anxiety and generalized anxiety disorder activate both the hypothalamic-pituitary-adrenal and the sympathoadrenal axes, panic attack causes major sympathetic activation, but has little effect on the hypothalamic-pituitary-adrenal axis.