Effects of long-term cocaine self-administration on brain resting-state functional connectivity in nonhuman primates.

Long-term cocaine use is associated with a variety of neural and behavioral deficits that impact daily function. This study was conducted to examine the effects of chronic cocaine self-administration on resting-state functional connectivity of the dorsal anterior cingulate (dACC) and putamen-two brain regions involved in cognitive function and motoric behavior-identified in a whole brain analysis. Six adult male squirrel monkeys self-administered cocaine (0.32 mg/kg/inj) over 140 sessions. Six additional monkeys that had not received any drug treatment for ~1.5 years served as drug-free controls. Resting-state fMRI imaging sessions at 9.4 Tesla were conducted under isoflurane anesthesia. Functional connectivity maps were derived using seed regions placed in the left dACC or putamen. Results show that cocaine maintained robust self-administration with an average total intake of 367 mg/kg (range: 299-424 mg/kg). In the cocaine group, functional connectivity between the dACC seed and regions primarily involved in motoric behavior was weaker, whereas connectivity between the dACC seed and areas implicated in reward and cognitive processing was stronger. In the putamen seed, weaker widespread connectivity was found between the putamen and other motor regions as well as with prefrontal areas that regulate higher-order executive function; stronger connectivity was found with reward-related regions. dACC connectivity was associated with total cocaine intake. These data indicate that functional connectivity between regions involved in motor, reward, and cognitive processing differed between subjects with recent histories of cocaine self-administration and controls; in dACC, connectivity appears to be related to cumulative cocaine dosage during chronic exposure.

Conditional Human Immunodeficiency Virus Transactivator of Transcription Protein Expression Induces Depression-like Effects and Oxidative Stress.

BACKGROUND: The prevalence of major depression in those with HIV/AIDS is substantially higher than in the general population. Mechanisms underlying this comorbidity are poorly understood. HIV-transactivator of transcription (Tat) protein, produced and excreted by HIV, could be involved. We determined whether conditional Tat protein expression in mice is sufficient to induce depression-like behaviors and oxidative stress. Further, as oxidative stress is associated with depression, we determined whether decreasing or increasing oxidative stress by administering methylsulfonylmethane (MSM) or diethylmaleate (DEM), respectively, altered depression-like behavior. METHODS: GT-tg bigenic mice received intraperitoneal saline or doxycycline (Dox, 25-100 mg/kg/day) to induce Tat expression. G-tg mice, which do not express Tat protein, also received Dox. Depression-like behavior was assessed with the tail suspension test (TST) and the two-bottle saccharin/water consumption task. Reactive oxygen/nitrogen species (ROS/RNS) were assessed ex vivo. Medial frontal cortex (MFC) oxidative stress and temperature were measured in vivo with 9.4-Tesla proton magnetic resonance spectroscopy (MRS). RESULTS: Tat expression increased TST immobility time in an exposure-dependent manner and reduced saccharin consumption. MSM decreased immobility time while DEM increased it in saline-treated GT-tg mice. Tat and MSM behavioral effects persisted for 28 days. Tat and DEM increased while MSM decreased ROS/RNS levels. Tat expression increased MFC glutathione levels and temperature. CONCLUSIONS: Tat expression induced rapid and enduring depression-like behaviors and oxidative stress. Increasing/decreasing oxidative stress increased/decreased, respectively, depression-like behavior. Thus, Tat produced by HIV may contribute to the high depression prevalence among those with HIV. Further, mitigation of oxidative stress could reduce depression severity.

Effects of Near-Infrared Light on Cerebral Bioenergetics Measured with Phosphorus Magnetic Resonance Spectroscopy.

OBJECTIVE: Cerebral photobiomodulation (PBM) improves mood and cognition. Cerebral metabolic enhancement is a mechanism proposed to underlie PBM effects. No PBM studies to date have applied phosphorus magnetic resonance spectroscopy (31P MRS), which can be used to assess metabolic intermediates such as phosphocreatine (PCr) and adenosine triphosphate, the latter of which is elevated by PBM. Accordingly, we used 9.4 Tesla 31P MRS to characterize effects of single and repeat cerebral PBM treatments on metabolism. PBM was delivered to healthy adult beagles in the form of transcranial laser treatment (TLT) at a wavelength of 808 nm, which passes safely through the skull and activates cytochrome C oxidase, a mitochondrial respiratory chain enzyme. METHODS: Isoflurane-anesthetized subjects (n = 4) underwent a baseline 31P MRS scan followed by TLT applied sequentially for 2 min each to anterior and posterior cranium midline locations, to irradiate the dorsal cortex. Subjects then underwent 31P MRS scans for 2 h to assess acute TLT effects. After 2 weeks of repeat TLT (3 times/week), subjects were scanned again with 31P MRS to characterize effects of repeat TLT. RESULTS: TLT did not induce acute 31P MRS changes over the course of 2 h in either scan session. However, after repeat TLT, the baseline PCr/ß-nucleoside triphosphate ratio was higher than the scan 1 baseline (p < 0.0001), an effect attributable to increased PCr level (p < 0.0001). CONCLUSIONS: Our findings are consistent with reports that bioenergetic effects of PBM can take several hours to evolve. Thus, in vivo 31P MRS may be useful for characterizing bioenergetic effects of PBM in brain and other tissues.

Striatal magnetic resonance spectroscopy abnormalities in young adult SAPAP3 knockout mice.

BACKGROUND: Obsessive compulsive disorder (OCD) is a debilitating condition with lifetime prevalence of 1-3%. OCD typically arises in youth but delays in diagnosis impede optimal treatment and developmental studies of the disorder. Research using genetically modified rodents may provide models of etiology that enable earlier detection and intervention. The SAPAP3 knockout (KO) transgenic mouse was developed as an animal model of OCD and related disorders (OCRD). KO mice exhibit compulsive self-grooming behavior analogous to behaviors found in people with OCRD. Striatal hyperactivity has been reported in these mice and in humans with OCD. METHODS: Striatal and medial frontal cortex 9.4 Tesla proton spectra were acquired from young adult SAPAP3 KO and wild-type control mice to determine whether KO mice have metabolic and neurochemical abnormalities. RESULTS: Young adult KO mice had lower striatal lactate (P=0.006) and glutathione (P=0.039) levels. Among all mice, striatal lactate and glutathione levels were associated (R=0.73, P=0.007). We found no group differences in medial frontal cortex metabolites. At the age range studied, only 1 of 8 KO mice had skin lesions indicative of severe compulsive grooming. CONCLUSION: Young adult SAPAP3 KO mice have striatal but not medial frontal cortex MRS abnormalities that may reflect striatal hypermetabolism accompanied by oxidative stress. These abnormalities typically preceded the onset of severe compulsive grooming. Our findings are consistent with striatal hypermetabolism in OCD. Together, these results suggest that striatal MRS measures of lactate or glutathione might be useful biomarkers for early detection of risk for developing compulsive behavior disorders.

Cocaine-conditioned odor cues without chronic exposure: Implications for the development of addiction vulnerability.

Adolescents are highly vulnerable to addiction and are four times more likely to become addicted at first exposure than at any other age. The dopamine D1 receptor, which is typically overexpressed in the normal adolescent prefrontal cortex, is involved in drug cue responses and is associated with relapse in animal models. In human drug addicts, imaging methods have detected increased activation in response to drug cues in reward- and habit-associated brain regions. These same methods can be applied more quantitatively to rodent models. Here, changes in neuronal activation in response to cocaine-conditioned cues were observed using functional magnetic resonance imaging in juvenile rats that were made to over-express either D1 receptors or green fluorescent protein by viral-mediated transduction. Reduced activation was observed in the amygdala and dopamine cell body regions in the low cue-preferring/control juvenile rats in response to cocaine cues. In contrast, increased activation was observed in the dorsal striatum, nucleus accumbens, prefrontal cortex, and dopamine cell bodies in high cue-preferring/D1 juveniles. The increase in cue salience that is mediated by increased D1 receptor density, rather than excessive cocaine experience, appears to underlie the transition from aversion to reward in cue-induced neural response and may form the basis for habit-forming vulnerability.

In vivo magnetic resonance studies reveal neuroanatomical and neurochemical abnormalities in the serine racemase knockout mouse model of schizophrenia.

BACKGROUND: Decreased availability of the N-methyl-D-aspartate receptor (NMDAR) co-agonist D-serine is thought to promote NMDAR hypofunction and contribute to the pathophysiology of schizophrenia, including neuroanatomical abnormalities, such as cortical atrophy and ventricular enlargement, and neurochemical abnormalities, such as aberrant glutamate and γ-aminobutyric acid (GABA) signaling. It is thought that these abnormalities directly relate to the negative symptoms and cognitive impairments that are hallmarks of the disorder. Because of the genetic complexity of schizophrenia, animal models of the disorder are extremely valuable for the study of genetically predisposing factors. Our laboratory developed a transgenic mouse model lacking serine racemase (SR), the synthetic enzyme of d-serine, polymorphisms of which are associated with schizophrenia. Null mutants (SR-/-) exhibit NMDAR hypofunction and cognitive impairments. We used 9.4 T magnetic resonance imaging (MRI) and proton spectroscopy (MRS) to compare in vivo brain structure and neurochemistry in wildtype (WT) and SR-/- mice. METHODS: Mice were anesthetized with isoflurane for MRI and MRS scans. RESULTS: Compared to WT controls, SR-/- mice exhibited 23% larger ventricular volumes (p<0.05). Additionally, in a medial frontal cortex voxel (15 µl), SR-/- mice exhibited significantly higher glutamate/water (12%, t=1.83, p<0.05) and GABA/water (72%, t=4.10, p<0.001) ratios. CONCLUSIONS: Collectively, these data demonstrate in vivo neuroanatomical and neurochemical abnormalities in the SR-/- mouse comparable to those previously reported in humans with schizophrenia.

Xenon impairs reconsolidation of fear memories in a rat model of post-traumatic stress disorder (PTSD).

Xenon (Xe) is a noble gas that has been developed for use in people as an inhalational anesthestic and a diagnostic imaging agent. Xe inhibits glutamatergic N-methyl-D-aspartate (NMDA) receptors involved in learning and memory and can affect synaptic plasticity in the amygdala and hippocampus, two brain areas known to play a role in fear conditioning models of post-traumatic stress disorder (PTSD). Because glutamate receptors also have been shown to play a role in fear memory reconsolidation--a state in which recalled memories become susceptible to modification--we examined whether Xe administered after fear memory reactivation could affect subsequent expression of fear-like behavior (freezing) in rats. Male Sprague-Dawley rats were trained for contextual and cued fear conditioning and the effects of inhaled Xe (25%, 1 hr) on fear memory reconsolidation were tested using conditioned freezing measured days or weeks after reactivation/Xe administration. Xe administration immediately after fear memory reactivation significantly reduced conditioned freezing when tested 48 h, 96 h or 18 d after reactivation/Xe administration. Xe did not affect freezing when treatment was delayed until 2 h after reactivation or when administered in the absence of fear memory reactivation. These data suggest that Xe substantially and persistently inhibits memory reconsolidation in a reactivation and time-dependent manner, that it could be used as a new research tool to characterize reconsolidation and other memory processes, and that it could be developed to treat people with PTSD and other disorders related to emotional memory.

A method for conducting functional MRI studies in alert nonhuman primates: initial results with opioid agonists in male cynomolgus monkeys.

Functional MRI (fMRI) has emerged as a powerful technique for assessing neural effects of psychoactive drugs and other stimuli. Several experimental approaches have been developed to use fMRI in anesthetized and awake animal subjects, each of which has its advantages and complexities. We sought to assess whether one particular method to scan alert postanesthetized animals can be used to assess fMRI effects of opioid agonists. To date, the use of fMRI as a method to compare pharmacological effects of opioid drugs has been limited. Such studies are important because mu and kappa opioid receptor agonists produce distinct profiles of behavioral effects related both to clinically desirable endpoints (e.g., analgesia) and to undesirable effects (e.g., abuse potential). This study sought to determine whether we could use our fMRI approach to compare acute effects of behaviorally equipotent (3.2 µg/kg) intravenous doses of fentanyl and U69,593 (doses that do not affect cardiorespiratory parameters). Scans were acquired in alert male cynomolgus macaques acclimated to undergo fMRI scans under restraint, absent excessive stress hormone increases. These opioid agonists activated bilateral striatal and nucleus accumbens regions of interest. At the dose tested, U69,593 induced greater left nucleus accumbens BOLD activation than fentanyl, while fentanyl activated left dorsal caudate nucleus more than U69,593. Our results suggest that our fMRI approach could be informative for comparing effects of opioid agonists.

Brain structural abnormalities in Doberman pinschers with canine compulsive disorder.

Obsessive compulsive disorder (OCD) is a debilitating condition, the etiology of which is poorly understood, in part because it often remains undiagnosed/untreated for a decade or more. Characterizing the etiology of compulsive disorders in animal models may facilitate earlier diagnosis and intervention. Doberman pinschers have a high prevalence of an analogous behavioral disorder termed canine compulsive disorder (CCD), which in many cases responds to treatments used for OCD. Thus, studies of CCD may help elucidate the etiology of compulsive disorders. We compared brain structure in Dobermans with CCD (N=8) and unaffected controls (N=8) to determine whether CCD is associated with structural abnormalities comparable to those reported in humans with OCD. We obtained 3 Tesla magnetic resonance structural and diffusion images from anesthetized Dobermans and subjected images to segmentation, voxel based morphometry, and diffusion tensor analyses. CCD dogs exhibited higher total brain and gray matter volumes and lower dorsal anterior cingulate cortex and right anterior insula gray matter densities. CCD dogs also had higher fractional anisotropy in the splenium of the corpus callosum, the degree of which correlated with the severity of the behavioral phenotype. Together, these findings suggest that CCD is associated with structural abnormalities paralleling those identified in humans with OCD. Accordingly, the CCD model, which has a number of advantages over other animal models of OCD, may assist in establishing the neuroanatomical basis for and etiology of compulsive disorders, which could lead to earlier diagnosis of and new treatments for humans and animals with these disorders.

Positive reinforcement training in squirrel monkeys using clicker training.

Nonhuman primates in research environments experience regular stressors that have the potential to alter physiology and brain function, which in turn can confound some types of research studies. Operant conditioning techniques such as positive reinforcement training (PRT), which teaches animals to voluntarily perform desired behaviors, can be applied to improve behavior and reactivity. PRT has been used to train rhesus macaques, marmosets, and several other nonhuman primate species. To our knowledge, the method has yet to be used to train squirrel monkeys to perform complex tasks. Accordingly, we sought to establish whether PRT, utilizing a hand-box clicker (which emits a click sound that acts as the conditioned reinforcer), could be used to train adult male squirrel monkeys (Saimiri boliviensis, N = 14). We developed and implemented a training regimen to elicit voluntary participation in routine husbandry, animal transport, and injection procedures. Our secondary goal was to quantify the training time needed to achieve positive results. Squirrel monkeys readily learned the connection between the conditioned reinforcer (the clicker) and the positive reinforcer (food). They rapidly developed proficiency on four tasks of increasing difficulty: target touching, hand sitting, restraint training, and injection training. All subjects mastered target touching behavior within 2 weeks. Ten of 14 subjects (71%) mastered all tasks in 59.2 ± 2.6 days (range: 50-70 days). In trained subjects, it now takes about 1.25 min per monkey to weigh and administer an intramuscular injection, one-third of the time it took before training. From these data, we conclude that clicker box PRT can be successfully learned by a majority of squirrel monkeys within 2 months and that trained subjects can be managed more efficiently. These findings warrant future studies to determine whether PRT may be useful in reducing stress-induced experimental confounds in studies involving squirrel monkeys.

Chronic cocaine exposure induces putamen glutamate and glutamine metabolite abnormalities in squirrel monkeys.

RATIONALE: Chronic cocaine exposure has been associated with progressive brain structural and functional changes. Clarifying mechanisms underlying cocaine's progressive brain effects may help in the development of effective cocaine abuse treatments. OBJECTIVES: We used a controlled squirrel monkey model of chronic cocaine exposure (45 mg/kg/week for 9 months) combined with ultra-high magnetic field (9.4 T) proton magnetic resonance spectroscopy to prospectively measure putamen metabolite changes. METHODS: Proton metabolites were measured with a STEAM sequence, quantified with LCModel using a simulated basis set, and expressed as metabolite/total creatine (tCr) ratios. RESULTS: We found cocaine-induced time-dependent changes in putamen glutamate/tCr and glutamine/tCr metabolite ratios suggestive of altered glutamate compartmentalization, neurotransmission, and metabolism. By contrast, saline-treated monkeys exhibited no metabolite changes over time. The time course of cocaine-induced metabolite abnormalities we detected is consistent with the apparent time course of glutamate abnormalities identified in a cross-sectional study in human cocaine users, as well as with microdialysis findings in rodent models of repeated cocaine exposure. CONCLUSIONS: Together, these findings suggests that this squirrel monkey model may be useful for characterizing glutamatergic changes associated with cocaine exposure and for determining efficacies of treatments designed to mitigate cocaine-induced glutamatergic system dysfunction.