Deprenyl reduces inflammation during acute SIV infection.

In the era of antiretroviral therapy, inflammation is a central factor in numerous HIV-associated comorbidities, such as cardiovascular disease, cognitive impairment, and neuropsychiatric disorders. This highlights the value of developing therapeutics that both reduce HIV-associated inflammation and treat associated comorbidities. Previous research on monoamine oxidase inhibitors (MAOIs) suggests this class of drugs has anti-inflammatory properties in addition to neuropsychiatric effects. Therefore, we examined the impact of deprenyl, an MAOI, on SIV-associated inflammation during acute SIV infection using the rhesus macaque model of HIV infection. Our results show deprenyl decreased both peripheral and CNS inflammation but had no effect on viral load in either the periphery or CNS. These data show that the MAOI deprenyl may have broad anti-inflammatory effects when given during the acute stage of SIV infection, suggesting more research into the anti-inflammatory effects of this drug could result in a beneficial adjuvant for antiretroviral therapy.

Dopamine transporter function fluctuates across sleep/wake state: potential impact for addiction.

The dopamine transporter (DAT) has been implicated in a variety of arousal-related processes including the regulation of motor activity, learning, motivated behavior, psychostimulant abuse, and, more recently, sleep/wake state. We previously demonstrated that DAT uptake regulates fluctuations in extracellular dopamine (DA) in the striatum across the light/dark cycle with DA levels at their highest during the dark phase and lowest during the light phase. Despite this evidence, whether fluctuations in DA uptake across the light/dark cycle are associated with changes in sleep/wake has not been tested. To address this, we employed a combination of sleep/wake recordings, fast scan cyclic voltammetry, and western blotting to examine whether sleep/wake state and/or light/dark phase impact DA terminal neurotransmission in male rats. Further, we assessed whether variations in plasma membrane DAT levels and/or phosphorylation of the threonine 53 site on the DAT accounts for fluctuations in DA neurotransmission. Given the extensive evidence indicating that psychostimulants increase DA through interactions with the DAT, we also examined to what degree the effects of cocaine at inhibiting the DAT vary across sleep/wake state. Results demonstrated a significant association between individual sleep/wake states and DA terminal neurotransmission, with higher DA uptake rate, increased phosphorylation of the DAT, and enhanced cocaine potency observed after periods of sleep. These findings suggest that sleep/wake state influences DA neurotransmission in a manner that is likely to impact a host of DA-dependent processes including a variety of neuropsychiatric disorders.

Hypocretin receptor 1 blockade produces bimodal modulation of cocaine-associated mesolimbic dopamine signaling.

RATIONALE: Cocaine addiction is a chronic psychiatric disorder characterized by pathological motivation to obtain cocaine and behavioral and neurochemical hypersensitivity to cocaine-associated cues. These features of cocaine addiction are thought to be driven by aberrant phasic dopamine signaling. We previously demonstrated that blockade of the hypocretin receptor 1 (HCRTr1) attenuates cocaine self-administration and reduces cocaine-induced enhancement of dopamine signaling. Despite this evidence, the effects of HCRTr1 blockade on endogenous phasic dopamine release are unknown. OBJECTIVE: In the current studies, we assessed whether blockade of HCRTr1 alters spontaneous and cue-evoked dopamine release in the nucleus accumbens core of freely moving rats. METHODS: We first validated the behavioral and neurochemical effects of the novel, highly selective, HCRTr1 antagonist RTIOX-276 using cocaine self-administration and fast-scan cyclic voltammetry (FSCV) in anesthetized rats. We then used FSCV in freely moving rats to examine whether RTIOX-276 impacts spontaneous and cue-evoked dopamine release. Finally, we used ex vivo slice FSCV to determine whether the effects of RTIOX-276 on dopamine signaling involve dopamine terminal adaptations. RESULTS: Doses of RTIOX-276 that attenuate the motivation for cocaine reduce spontaneous dopamine transient amplitude and cue-evoked dopamine release. Further, these doses attenuated cocaine-induced dopamine uptake inhibition at the level of dopamine terminals. CONCLUSION: Our results provide support for the standing hypothesis that HCRTr1 blockade suppresses endogenous phasic dopamine signals, likely via actions at dopamine cell bodies. These results also elucidate a second process through which HCRTr1 blockade attenuates the effects of cocaine by reducing cocaine sensitivity at dopamine terminals.

Low and high affinity dopamine transporter inhibitors block dopamine uptake within 5 sec of intravenous injection.

Extensive evidence suggests that the reinforcing effects of cocaine involve inhibition of dopamine transporters (DAT) and subsequent increases in dopamine (DA) levels in the striatum. We have previously reported that cocaine inhibits the DAT within 4-5 s of i.v. injection, matching the temporal profile of the behavioral and subjective effects of cocaine. Intravenous injection of GBR-12909, a high affinity, long-acting DAT inhibitor, also inhibits DA uptake within 5 s. Given that high affinity, long-acting drugs are considered to have relatively low abuse potential, we found it intriguing that GBR-12909 had an onset profile similar to that of cocaine. To further explore the onset kinetics of both low and high affinity DAT inhibitors, we examined the effects of i.v. cocaine (1.5 mg/kg), methylphenidate (1.5 mg/kg), nomifensine (1.5 mg/kg), GBR-12909 (1.5 mg/kg), PTT (0.5 mg/kg), and WF23 (0.5 mg/kg) on electrically-evoked DA release and uptake in the nucleus accumbens core. Results indicate that all of the DAT inhibitors significantly inhibited DA uptake within 5 s of injection. However, the timing of peak uptake inhibition varied greatly between the low and high affinity uptake inhibitors. Uptake inhibition following cocaine, methylphenidate, and nomifensine peaked 30 s following injection. In contrast, peak effects for GBR-12909, PTT, and WF23 occurred between 20 and 60 min following injection. These observations suggest that the initial onset for i.v. DAT inhibitors is extremely rapid and does not appear to be dictated by a drug's affinity.

Mechanisms underlying cognitive enhancement and reversal of cognitive deficits in nonhuman primates by the ampakine CX717.

RATIONALE: Performance of cognitive tasks in nonhuman primates (NHPs) requires specific brain regions to make decisions under different degrees of difficulty or "cognitive load." OBJECTIVE: Local cerebral metabolic activity ([18F]FDG PET imaging) in dorsolateral prefrontal cortex (DLPFC), medial temporal lobe (MTL), and dorsal striatum (DStr) is examined in NHPs performing a delayed-match-to-sample (DMS) task with variable degrees of cognitive load. MATERIALS AND METHODS: Correlations between cognitive load and degree of brain metabolic activity were obtained with respect to the influence of the ampakine CX717 (Cortex Pharmaceuticals), using brain imaging and recordings of neuronal activity in NHPs and measures of intracellular calcium release in rat hippocampal slices. RESULTS: Activation of DLPFC, MTL, and DStr reflected changes in performance related to cognitive load within the DMS task and were engaged primarily on high load trials. Similar increased activation patterns and improved performance were also observed following administration of CX717. Sleep deprivation in NHPs produced impaired performance and reductions in brain activation which was reversed by CX717. One potential basis for this facilitation of cognition by CX717 was increased firing of task-specific hippocampal cells. Synaptic mechanisms affected by CX717 were examined in rat hippocampal slices which showed that N-methyl-D-aspartic acid-mediated release of intracellular calcium was reduced in slices from sleep-deprived rats and reversed by application of CX717 to the bathing medium. CONCLUSIONS: The findings provide insight into how cognition is enhanced by CX717 in terms of brain, and underlying neural, processes that are activated on high vs. low cognitive load trials.

Short-acting cocaine and long-acting GBR-12909 both elicit rapid dopamine uptake inhibition following intravenous delivery.

The rewarding effects of cocaine have been reported to occur within seconds of administration. Extensive evidence suggests that these actions involve the ability of cocaine to inhibit the dopamine (DA) transporter. We recently showed that 1.5 mg/kg i.v. cocaine inhibits DA uptake within 5 s. Despite this evidence, there remains a lack of consensus regarding how quickly i.v. cocaine and other DA uptake inhibitors elicit DA uptake inhibition. The current studies sought to better characterize the onset of cocaine-induced DA uptake inhibition and to compare these effects to those obtained with the high-affinity, long-acting DA transporter inhibitor, GBR-12909 (1-(2-bis(4-fluorphenyl)-methoxy)-ethyl)-4-(3-phenyl-propyl)piperazine). Using in vivo fast scan cyclic voltammetry, we showed that i.v. cocaine (0.75, 1.5, and 3.0 mg/kg) significantly inhibited DA uptake in the nucleus accumbens of anesthetized rats within 5 s. DA uptake inhibition peaked at 30 s and returned to baseline levels in approximately 1 h. The effects of cocaine were dose-dependent, with the 3.0 mg/kg dose producing greater uptake inhibition at the early time points and exhibiting a longer latency to return to baseline. Further, the blood-brain barrier impermeant cocaine-methiodide had no effect on DA uptake or peak height, indicating that the generalized peripheral effects of cocaine do not contribute to the CNS alterations measured here. Finally, we show that GBR-12909 (0.75, 1.5, and 3.0 mg/kg) also significantly inhibited DA uptake within 5 s post-injection, although the peak effect and return to baseline were markedly delayed compared with cocaine, particularly at the highest dose. Combined, these observations indicate that the central effects of dopamine uptake inhibitors occur extremely rapidly following i.v. drug delivery.

Concomitant loss of dynorphin, NARP, and orexin in narcolepsy.

BACKGROUND: Narcolepsy with cataplexy is associated with a loss of orexin/hypocretin. It is speculated that an autoimmune process kills the orexin-producing neurons, but these cells may survive yet fail to produce orexin. OBJECTIVE: To examine whether other markers of the orexin neurons are lost in narcolepsy with cataplexy. METHODS: We used immunohistochemistry and in situ hybridization to examine the expression of orexin, neuronal activity-regulated pentraxin (NARP), and prodynorphin in hypothalami from five control and two narcoleptic individuals. RESULTS: In the control hypothalami, at least 80% of the orexin-producing neurons also contained prodynorphin mRNA and NARP. In the patients with narcolepsy, the number of cells producing these markers was reduced to about 5 to 10% of normal. CONCLUSIONS: Narcolepsy with cataplexy is likely caused by a loss of the orexin-producing neurons. In addition, loss of dynorphin and neuronal activity-regulated pentraxin may contribute to the symptoms of narcolepsy.

Fos immunoreactivity in hypocretin-synthesizing and hypocretin-1 receptor-expressing neurons: effects of diurnal and nocturnal spontaneous waking, stress and hypocretin-1 administration.

Hypocretin/orexin modulates sleep-wake state via actions across multiple terminal fields. Within waking, hypocretin may also participate in high-arousal processes, including those associated with stress. The current studies examined the extent to which alterations in neuronal activity, as measured by Fos immunoreactivity, occur within both hypocretin-synthesizing and hypocretin-1 receptor-expressing neurons across varying behavioral state/environmental conditions associated with varying levels of waking and arousal. Double-label immunohistochemistry was used to visualize Fos and either prepro-hypocretin in the lateral hypothalamus or hypocretin-1 receptors in the locus coeruleus and select basal forebrain regions involved in the regulation of behavioral state/arousal. Animals were tested under the following conditions: 1). diurnal sleeping; 2). diurnal spontaneous waking; 3). nocturnal spontaneous waking; and 4). high-arousal waking (diurnal novelty-stress). Additionally, the effects of hypocretin-1 administration (0.07 and 0.7 nmol) on levels of Fos were examined within these two neuronal populations. Time spent awake, scored for the 90-min preceding perfusion, was largely comparable in diurnal spontaneous waking, nocturnal spontaneous waking and high-arousal waking. Nocturnal spontaneous waking and high-arousal waking, but not diurnal spontaneous waking, were associated with increased levels of Fos within hypocretin-synthesizing neurons, relative to diurnal sleeping. Within hypocretin-1 receptor-expressing neurons, only high-arousal waking was associated with increased levels of Fos. Hypocretin-1 administration dose-dependently increased levels of Fos within hypocretin-1 receptor-expressing neurons to levels comparable to, or exceeding, levels observed in high-arousal waking. Combined, these observations support the hypothesis that hypocretin neuronal activity varies across the circadian cycle. Additionally, these data suggest that waking per se may not be associated with increased hypocretin neurotransmission. In contrast, high-arousal states, including stress, appear to be associated with substantially higher rates of hypocretin neurotransmission. Finally, these studies provide further evidence indicating coordinated actions of hypocretin across a variety of arousal-related basal forebrain and brainstem regions in the behavioral state modulatory actions of this peptide system.

Additive wake-promoting actions of medial basal forebrain noradrenergic alpha1- and beta-receptor stimulation.

The locus coeruleus-noradrenergic system exerts an activating influence on forebrain neuronal and behavioral activity states, in part through the actions of noradrenergic beta-receptors in the medial septal (MS) and medial preoptic (MPOA) areas. MPOA alpha1-receptors exert similar wake-promoting actions. The current study examines the influence of alpha1-receptors located within MS on sleep-wake state. In addition, the extent to which alpha1- and beta-receptors located within MS and MPOA interact in the modulation of behavioral state was investigated by examining the effects of individual or combined infusion of alpha1- and beta-agonists into these regions. Results show that alpha1-receptors located within MS exert wake-promoting actions. Within both MS and MPOA, additive wake-promoting actions were observed with alpha1- and beta-receptor stimulation, the sum of which contributes to the overall arousal state of the animal.

Prolactin-releasing peptide (PrRP) promotes awakening and suppresses absence seizures.

Prolactin releasing peptide (PrRP) is a recently identified neuropeptide that stimulates prolactin release from pituitary cells. The presence of its receptor outside the hypothalamic-pituitary axis suggests that it may have other functions. We present here evidence that PrRP can modulate the activity of the reticular thalamic nucleus, a brain region with prominent PrRP receptor expression that is critical for sleep regulation and the formation of non-convulsive absence seizures. Intracerebroventricular injection of PrRP (1-10 nmol) into sleeping animals significantly suppresses sleep oscillations and promotes rapid and prolonged awakening. Higher concentrations of PrRP (10-100 nmol) similarly suppress spike wave discharges seen during absence seizures in genetic absence epilepsy rats from Strasbourg, an animal model for this disorder. In concordance with these findings, PrRP suppressed evoked oscillatory burst activity in reticular thalamic slices in vitro. These results indicate that PrRP modulates reticular thalamic function and that activation of its receptor provides a new target for therapies directed at sleep disorders and absence seizures.

Wake-promoting and sleep-suppressing actions of hypocretin (orexin): basal forebrain sites of action.

The hypocretins (orexins) are a newly identified peptide family comprised of two peptides, hypocretin-1 and hypocretin-2. Recent observations suggest an involvement of these peptides in the regulation of behavioral state. For example, these peptides are found in a variety of brain regions associated with the regulation of forebrain neuronal and behavioral activity states. Furthermore, when infused into the lateral ventricles in awake animals, hypocretin-1 elicits increased duration of waking beyond that observed in vehicle-treated animals. Previous studies have been limited to an examination of the sleep-wake effects of hypocretin-1 in awake animals. Currently, the sleep-wake effects of hypocretin-2 and the extent to which hypocretins can initiate waking in the sleeping animal remain unclear. To better characterize the wake-promoting actions of the hypocretins, the current studies examined the sleep-wake effects of varying doses (0.007, 0.07 and 0.7 nmol) of hypocretin-1 and hypocretin-2 when administered into sleeping rats (e.g. remote-controlled infusions). Infusions of hypocretin-1 and hypocretin-2 into the lateral ventricles elicited a short latency (0.7 nmol hypocretin-1; 93+/-30 s from the start of the 120-s infusion) increase in electroencephalographic, electromyographic, and behavioral indices of waking. These infusions also produced substantial decreases in slow-wave and rapid-eye movement sleep. Hypocretin-1 was more potent than hypocretin-2 in these actions. Interestingly, hypocretin-1 infused into the fourth ventricle elicited less robust waking which occurred with a longer latency than infusions into the lateral ventricles. These latter observations suggest a forebrain site of action participates in hypocretin-1-induced waking. Within the forebrain, a variety of basal forebrain structures, including the medial preoptic area, the medial septal area and the substantia innominata, receive a moderate hypocretin innervation. Therefore, additional studies examined the sleep-wake effects of bilateral hypocretin-1 infusions into these basal forebrain structures. Robust increases in waking were observed following infusions into, but not outside, the medial septal area, the medial preoptic area and the substantia innominata. These results indicate a potentially prominent role of hypocretins in sleep-wake regulation via actions within certain basal forebrain structures and are consistent with studies indicating a prominent role of hypocretins in sleep/arousal disorders.

Synergistic sedative effects of noradrenergic alpha(1)- and beta-receptor blockade on forebrain electroencephalographic and behavioral indices.

The locus coeruleus-noradrenergic system exerts an activating influence on forebrain neuronal and behavioral activity states. For example, in the anesthetized rat, unilateral locus coeruleus stimulation elicits bilateral activation of forebrain electroencephalographic activity. Pretreatment with a noradrenergic beta-antagonist blocks this effect, suggesting that beta-receptors play a critical role in locus coeruleus-dependent activation of the forebrain. Consistent with this, stimulation of beta-receptors located in certain basal forebrain structures evokes sustained periods of alert waking in the unanesthetized rat. Similar forebrain and behavioral activating effects are observed with alpha(1)-receptor stimulation within these basal forebrain regions. To assess the extent to which alpha(1)- and beta-receptors contribute to the maintenance of behavioral and forebrain activation, we examined the electroencephalographic and behavioral effects of alpha(1)-, beta- and combined alpha(1)/beta-receptor blockade in the unanesthetized rat. Rats were treated individually or in combination with either varying doses of the alpha(1)-antagonist, prazosin (intraperitoneally), and/or the beta-antagonist, timolol (intracerebroventricularly). Thirty minutes following treatment, animals were placed in a mildly-arousing novel environment, which has been demonstrated previously to elicit activation of central noradrenergic systems and sustained waking in vehicle-treated controls. Behavior and electroencephalographic activity were recorded and later scored. Electroencephalographic activity was analysed using power spectrum analysis. The following were observed: (i) beta-receptor blockade alone does not alter behavioral or electroencephalographic indices of alert waking; (ii) alpha(1)-receptor blockade alone increases high-voltage spindle activity in cortical electroencephalographic activity that was associated with decreased behavioral activity; (iii) combined alpha(1)- and beta-receptor blockade elicits a substantial increase in slow-wave activity (0.33-2.0Hz), also in association with decreased behavioral activity. All of these effects were dependent on the dose administered and time following initiation of testing. These results indicate that the combined actions of alpha(1)- and beta-receptors exert distinct and synergistic actions on cortical neuronal activity patterns that are essential elements of alert waking.