Effects of Bedtime Dosing With Suvorexant and Zolpidem on Balance and Psychomotor Performance in Healthy Elderly Participants During the Night and in the Morning.

PURPOSE/BACKGROUND: This study was designed as an early assessment of the safety of the orexin receptor antagonist suvorexant, but also included exploratory assessments of balance and psychomotor performance that are the focus of this report. METHODS/PROCEDURES: This was a double-blind, randomized, 3-period, crossover, phase 1 study. Balance and psychomotor performance were evaluated during the night in 12 healthy elderly participants after bedtime administration of suvorexant 30 mg (a supratherapeutic dose), the GABAergic agonist zolpidem 5 mg (the recommended dose in the elderly), or placebo. Balance (body sway measured by platform stability) and psychomotor performance (measured by choice reaction time) were assessed predose and at 1.5, 4, and 8 hours postdose in each period. Memory (measured by word recall) was assessed predose and at 4 hours postdose. FINDINGS/RESULTS: At 1.5 hours after nighttime administration of each drug (the approximate time of their anticipated maximal plasma concentrations), both zolpidem and suvorexant increased body sway versus placebo, with a greater increase for zolpidem than suvorexant. Suvorexant increased choice reaction time compared with placebo or zolpidem at 1.5 hours. There were no treatment differences on body sway or choice reaction time at 4 or 8 hours, or on word recall at 4 hours. IMPLICATIONS/CONCLUSIONS: These exploratory data suggest that a 30-mg dose of suvorexant (supratherapeutic) and a 5-mg dose of zolpidem (recommended dose in the elderly) impaired balance at 1.5 hours in healthy elderly people, with potentially less impairment for suvorexant relative to zolpidem, but no treatment differences on body sway or psychomotor performance at 4 and 8 hours. Because of their exploratory nature, these findings and their clinical relevance, if any, require confirmation in a prospective study.

Dysregulation of behavioral and autonomic responses to emotional and social stimuli following bidirectional pharmacological manipulation of the basolateral amygdala in macaques.

The amygdala is a key component of the neural circuits mediating the processing and response to emotionally salient stimuli. Amygdala lesions dysregulate social interactions, responses to fearful stimuli, and autonomic functions. In rodents, the basolateral and central nuclei of the amygdala have divergent roles in behavioral control. However, few studies have selectively examined these nuclei in the primate brain. Moreover, the majority of non-human primate studies have employed lesions, which only allow for unidirectional manipulation of amygdala activity. Thus, the effects of amygdala disinhibition on behavior in the primate are unknown. To address this gap, we pharmacologically inhibited by muscimol or disinhibited by bicuculline methiodide the basolateral complex of the amygdala (BLA; lateral, basal, and accessory basal) in nine awake, behaving male rhesus macaques (Macaca mulatta). We examined the effects of amygdala manipulation on: (1) behavioral responses to taxidermy snakes and social stimuli, (2) food competition and social interaction in dyads, (3) autonomic arousal as measured by cardiovascular response, and (4) prepulse inhibition of the acoustic startle (PPI) response. All modalities were impacted by pharmacological inhibition and/or disinhibition. Amygdala inhibition decreased fear responses to snake stimuli, increased examination of social stimuli, reduced competitive reward-seeking in dominant animals, decreased heart rate, and increased PPI response. Amygdala disinhibition restored fearful response after habituation to snakes, reduced competitive reward-seeking behavior in dominant animals, and lowered heart rate. Thus, both hypoactivity and hyperactivity of the basolateral amygdala can lead to dysregulated behavior, suggesting that a narrow range of activity is necessary for normal functions.

MIDD0301 - A first-in-class anti-inflammatory asthma drug targets GABAA receptors without causing systemic immune suppression.

We report a 28-day repeat dose immunotoxicity evaluation of investigational drug MIDD0301, a novel oral asthma drug candidate that targets gamma amino butyric acid type A receptors (GABAA R) in the lung. The study design employed oral administration of mice twice daily throughout the study period with 100 mg/kg MIDD0301 mixed in peanut butter. Compound dosing did not reveal signs of general toxicity as determined by animal weight, organ weight or haematology. Peanut butter plus test drug (in addition to ad libitum standard rodent chow) did not affect weight gain in the adult mice, in contrast to weight loss in 5 mg/kg prednisone-treated mice. Spleen and thymus weights were unchanged in MIDD0301-treated mice, but prednisone significantly reduced the weight of those organs over the 28-day dosing. Similarly, no differences in spleen or thymus histology were observed following MIDD0301 treatment, but prednisone treatment induced morphological changes in the spleen. The number of small intestine Peyer's patches was not affected by MIDD0301 treatment, an important factor for orally administered drugs. Circulating lymphocyte, monocyte and granulocyte numbers were unchanged in the MIDD0301-treated animals, whereas differential lymphocyte numbers were reduced in prednisone-treated animals. MIDD0301 treatment did not alter IgG antibody responses to dinitrophenyl following dinitrophenyl-keyhole limpet haemocyanin immunization, indicating that systemic humoral immune function was not affected. Taken together, these studies show that repeated daily administration of MIDD0301 is safe and not associated with adverse immunotoxicological effects in mice.

Involvement of glutamatergic N-methyl-d-aspartate receptors in the expression of increased head-dipping behaviors in the hole-board tests of olfactory bulbectomized mice.

Olfactory bulbectomized (OB) mice produce agitated anxiety-like behaviors in the hole-board test, which was expressed by an increase in head-dipping counts and a decrease in head-dipping latencies. However, the associated mechanisms remain unclear. In the present study, MK-801 (10, 100µg/kg), a selective N-methyl-d-aspartate (NMDA) receptor antagonist, significantly and dose-dependently suppressed the increased head-dipping behaviors in OB mice, without affecting sham mice. Similar results were obtained with another selective NMDA receptor antagonist D-AP5 treatment in OB mice. On the other hand, muscimol, a selective aminobutyric acid type A (GABAA) receptor agonist produced no effects on these hyperemotional behaviors in OB mice at a dose (100µg/kg) that produced anxiolytic-like effects in sham mice. Interestingly, glutamine contents and glutamine/glutamate ratios were significantly increased in the amygdala and frontal cortex of OB mice compared to sham mice. Based on these results, we concluded that the glutamatergic NMDA receptors are involved in the expression of increased head-dipping behaviors in the hole-board tests of OB mice. Accordingly, the changes in glutamatergic transmission in frontal cortex and amygdala may play important roles in the expression of these abnormal behaviors in OB mice.

Hypothalamic Paraventricular and Arcuate Nuclei Contribute to Elevated Sympathetic Nerve Activity in Pregnant Rats: Roles of Neuropeptide Y and α-Melanocyte-Stimulating Hormone.

Pregnancy increases sympathetic nerve activity (SNA), but the mechanisms are unknown. Here, we investigated the contributions of the hypothalamic paraventricular and arcuate nuclei in α-chloralose-anesthetized pregnant and nonpregnant rats. Baseline arterial pressure (AP) was lower, and heart rate (HR), lumbar sympathetic activity, and splanchnic SNA were higher in pregnant rats compared with nonpregnant rats. Inhibition of the paraventricular nucleus via bilateral muscimol nanoinjections decreased AP and HR more in pregnant rats than in nonpregnant rats and decreased lumbar SNA only in pregnant rats. Similarly, after arcuate muscimol nanoninjections, the decreases in AP, HR, and lumbar, renal, and splanchnic sympathetic nerve activities were greater in pregnant rats than in nonpregnant rats. Major arcuate neuronal groups that project to the paraventricular nucleus express inhibitory neuropeptide Y (NPY) and excitatory α-melanocyte-stimulating hormone. Inhibition of paraventricular melanocortin 3/4 receptors with SHU9119 also decreased AP, HR, and lumbar SNA in pregnant rats but not in nonpregnant rats. Conversely, paraventricular nucleus NPY expression was reduced in pregnant animals, and although blockade of paraventricular NPY Y1 receptors increased AP, HR, and lumbar sympathetic activity in nonpregnant rats, it had no effects in pregnant rats. Yet, the sympathoinhibitory, depressor, and bradycardic effects of paraventricular NPY nanoinjections were similar between groups. In conclusion, the paraventricular and arcuate nuclei contribute to increased basal SNA during pregnancy, likely due in part to decreased tonic NPY inhibition and increased tonic α-melanocyte-stimulating hormone excitation of presympathetic neurons in the paraventricular nucleus.

γ-Aminobutyric acid neural signaling in the lateroanterior hypothalamus modulates aggressive behavior in adolescent anabolic/androgenic steroid-treated hamsters.

Male Syrian hamsters (Mesocricetus auratus) treated with anabolic/androgenic steroids (AAS) during adolescence (P27-P56) display highly escalated and mature forms of offensive aggression correlated with increased γ-aminobutyric acid (GABA) afferent development as well as decreased GABAA receptors in the lateroanterior hypothalamus (LAH) - an area of convergence for developmental and neuroplastic changes that underlie offensive aggressive behaviors in hamsters. This study investigated whether microinfusion of a GABAA receptor agonist (muscimol; 0.01-1.0 pmol/l) or antagonist (bicuculline; 0.04-4.0 pmol/l) directly into the LAH modulate adolescent AAS-induced offensive aggression. Activation of LAH GABAA receptors enhanced adolescent AAS-induced offensive aggression, beginning at the 0.1 pmol/l dose, when compared with AAS-treated animals injected with saline into the LAH. Importantly, GABAA receptor agonism within the LAH significantly increased the frequency of belly/rear attacks, while simultaneously decreasing the frequency of frontal attacks. These data identify a neuroanatomical locus where GABAA receptor activation functions to enhance aggression in adolescent AAS-treated animals, while also promoting the display of mature forms of aggression and suppressing juvenile play behaviors.

GABAergic excitation of vasopressin neurons: possible mechanism underlying sodium-dependent hypertension.

RATIONALE: Increased arginine-vasopressin (AVP) secretion is a key physiological response to hyperosmotic stress and may be part of the mechanism by which high-salt diets induce or exacerbate hypertension. OBJECTIVE: Using deoxycorticosterone acetate-salt hypertension model rats, we sought to test the hypothesis that changes in GABA(A) receptor-mediated inhibition in AVP-secreting magnocellular neurons contribute to the generation of Na(+)-dependent hypertension. METHODS AND RESULTS: In vitro gramicidin-perforated recordings in the paraventricular and supraoptic nuclei revealed that the GABAergic inhibition in AVP-secreting neurons was converted into excitation in this model, because of the depolarization of GABA equilibrium potential. Meanwhile, in vivo extracellular recordings in the supraoptic nuclei showed that the GABAergic baroreflexive inhibition of magnocellular neurons was transformed to excitation, so that baroreceptor activation may increase AVP release. The depolarizing GABA equilibrium potential shift in AVP-secreting neurons occurred progressively over weeks of deoxycorticosterone acetate-salt treatment along with gradual increases in plasma AVP and blood pressure. Furthermore, the shift was associated with changes in chloride transporter expression and partially reversed by bumetanide (Na(+)-K(+)-2Cl(-) cotransporter inhibitor). Intracerebroventricular bumetanide administration during deoxycorticosterone acetate-salt treatment hindered the development of hypertension and rise in plasma AVP level. Muscimol (GABA(A) agonist) microinjection into the supraoptic nuclei in hypertensive rats increased blood pressure, which was prevented by previous intravenous V1a AVP antagonist injection. CONCLUSIONS: We conclude that the inhibitory-to-excitatory switch of GABAA receptor-mediated transmission in AVP neurons contributes to the generation of Na(+)-dependent hypertension by increasing AVP release. We speculate that normalizing the GABA equilibrium potential may have some utility in treating Na(+)-dependent hypertension.

Functional role of the cerebellum in gamma-band synchronization of the sensory and motor cortices.

The cerebellum is an essential structure for the control of movement. It sends abundant ascending projections to the cerebral cortex via the thalamus, but its contribution to cortical activity remains largely unknown. Here we studied its influence on cortical neuronal activity in freely moving rats. We demonstrate an excitatory action of the cerebellum on the motor thalamus and the motor cortex. We also show that cerebellar inactivation disrupts the gamma-band coherence of local field potential between the sensory and motor cortices during whisking. In contrast, phase locking of neuronal activities to local gamma oscillations was preserved in the sensory and motor cortices by cerebellar inactivation. These results indicate that the cerebellum contributes to coordinated sensorimotor cortical activities during motor activation and thus participates in the multiregional cortical processing of information.

Mechanisms and time course of vocal learning and consolidation in the adult songbird.

In songbirds, the basal ganglia outflow nucleus LMAN is a cortical analog that is required for several forms of song plasticity and learning. Moreover, in adults, inactivating LMAN can reverse the initial expression of learning driven via aversive reinforcement. In the present study, we investigated how LMAN contributes to both reinforcement-driven learning and a self-driven recovery process in adult Bengalese finches. We first drove changes in the fundamental frequency of targeted song syllables and compared the effects of inactivating LMAN with the effects of interfering with N-methyl-d-aspartate (NMDA) receptor-dependent transmission from LMAN to one of its principal targets, the song premotor nucleus RA. Inactivating LMAN and blocking NMDA receptors in RA caused indistinguishable reversions in the expression of learning, indicating that LMAN contributes to learning through NMDA receptor-mediated glutamatergic transmission to RA. We next assessed how LMAN's role evolves over time by maintaining learned changes to song while periodically inactivating LMAN. The expression of learning consolidated to become LMAN independent over multiple days, indicating that this form of consolidation is not completed over one night, as previously suggested, and instead may occur gradually during singing. Subsequent cessation of reinforcement was followed by a gradual self-driven recovery of original song structure, indicating that consolidation does not correspond with the lasting retention of changes to song. Finally, for self-driven recovery, as for reinforcement-driven learning, LMAN was required for the expression of initial, but not later, changes to song. Our results indicate that NMDA receptor-dependent transmission from LMAN to RA plays an essential role in the initial expression of two distinct forms of vocal learning and that this role gradually wanes over a multiday process of consolidation. The results support an emerging view that cortical-basal ganglia circuits can direct the initial expression of learning via top-down influences on primary motor circuitry.

MRK-409 (MK-0343), a GABAA receptor subtype-selective partial agonist, is a non-sedating anxiolytic in preclinical species but causes sedation in humans.

MRK-409 binds to α1-, α2-, α3- and α5-containing human recombinant GABA(A) receptors with comparable high affinity (0.21-0.40 nM). However, MRK-409 has greater agonist efficacy at the α3 compared with α1 subtypes (respective efficacies relative to the full agonist chlordiazepoxide of 0.45 and 0.18). This compound readily penetrates the brain in rats and occupies the benzodiazepine site of GABA(A) receptors, measured using an in vivo [(3)H]flumazenil binding assay, with an Occ(50) of 2.2 mg/kg p.o. and a corresponding plasma EC(50) of 115 ng/mL. Behaviourally, the α3-preferring agonist efficacy profile of MRK-409 produced anxiolytic-like activity in rodent and primate unconditioned and conditioned models of anxiety with minimum effective doses corresponding to occupancies, depending on the particular model, ranging from ∼35% to 65% yet there were minimal overt signs of sedation at occupancies greater than 90%. In humans, however, safety and tolerability studies showed that there was pronounced sedation at a dose of 2 mg, resulting in a maximal tolerated dose of 1 mg. This 2 mg dose corresponded to a C(max) plasma concentration of 28 ng/mL, which, based on the rodent plasma EC(50) for occupancy of 115 ng/mL, suggested that sedation in humans occurs at low levels of occupancy. This was confirmed in human positron emission tomography studies, in which [(11)C]flumazenil uptake following a single dose of 1 mg MRK-409 was comparable to that of placebo, indicating that occupancy of GABA(A) receptor benzodiazepine binding sites by MRK-409 was below the limits of detection (i.e. <10%). Taken together, these data show that MRK-409 causes sedation in humans at a dose (2 mg) corresponding to levels of occupancy considerably less than those predicted from rodent models to be required for anxiolytic efficacy (∼35-65%). Thus, the preclinical non-sedating anxiolytic profile of MRK-409 did not translate into humans and further development of this compound was halted.