Matthew P. Galloway

FACILITATION OF DOPAMINE RELEASE IN VIVO BY SEROTONIN AGONISTS : STUDIES WITH MICRODIALYSIS

Using microdialysis, changes in extraneuronal levels of dopamine (DA), and the metabolites of DA and serotonin (5-HT), were monitored concurrent with perfusion of 5-HT1 agonists into the anterior striata of anesthetized rats. Perfusion of 5-HT facilitated DA release in a dose dependent manner, and to a greater extent than any other agonist tested. Extraneuronal DA levels increased 34% with perfusion of 0.04 nmol 5-HT and 18-fold with perfusion of 4.0 nmol 5-HT. Perfusion with multiple doses of either 1-(m-chlorophenyl)piperazine (m-CPP) or trifluoromethylphenylpiperazine (TFMPP) also resulted in a dose-dependent facilitation of DA release with a 40% increase in extracellular DA produced by either 0.4 nmol m-CPP or 10.0 nmol TFMPP. A 50-fold increase in DA followed 40.0 nmol m-CPP, while 160 nmol TFMPP enhanced DA 11-fold. Local application of either 5-methoxy-3(1,2,3,6-tetrahydro-4-pyridinyl)-1H indole succinate (RU24969) or 8-hydroxy-2-(di-n-propylamino)tetralin hydrobromide (8-OH-DPAT) (2.0 nmol perfused over 20 min) increased extracellular DA by 300 and 40%, respectively. RU24969 (2.0 nmol) also facilitated DA release following systemic pretreatment with 8-OH-DPAT (100 micrograms/kg). Perfusion with fenfluramine to release endogenous 5-HT also increased extraneuronal DA in a dose-dependent manner, and this facilitation was prevented by pretreatment with the 5-HT reuptake inhibitor fluoxetine. The facilitation of DA release by 0.4 nmol 5-HT was reduced by pretreatment with the 5-HT1 antagonist pindolol (4.0 nmol). These results suggest that serotonergic innervation of the anterior striatum may exert a facilitatory influence on DA release.

Repeated Unpredictable Stress and Antidepressants Differentially Regulate Expression of the Bcl-2 Family of Apoptotic Genes in Rat Cortical, Hippocampal, and Limbic Brain Structures

Apoptosis has been proposed as a contributing cellular mechanism to the structural alterations that have been observed in stress-related mood disorders. Antidepressants, on the other hand, are hypothesized to exert trophic and/or neuroprotective actions. The present study examined the regulation of the major antiapoptotic (Bcl-2, Bcl-xl) and proapoptotic (Bax) genes by repeated unpredictable stress (an animal model of depression) and antidepressant treatments (ADT). In adult rats, exposure to unpredictable stress reduced Bcl-2 mRNA levels in the central nucleus of the amygdala (CeA), cingulate (Cg), and frontal (Fr) cortices. Bcl-xl mRNA was significantly decreased in hippocampal subfields. In contrast, chronic administration of clinically effective antidepressants from four different classes, ie fluoxetine, reboxetine, tranylcypromine, and electroconvulsive seizures (ECS) upregulated Bcl-2 mRNA expression in the Cg, Fr, and CeA. Reboxetine, tranylcypromine, and ECS selectively increased Bcl-xl, but not Bcl-2 mRNA expression in the hippocampus. Chemical ADT but not ECS, robustly enhanced Bcl-2 expression in the medial amygdaloid nucleus and ventromedial hypothalamus. Fluoxetine did not influence Bcl-xl expression in the hippocampus, but it was the only ADT that decreased Bax expression in this region. In the CeA, again in direct contrast to the stress effects, exposure to all classes of ADTs significantly increased Bcl-2 mRNA. The selective regulation of Bcl-xl and Bax in hippocampal subfields and of Bcl-2 in the Cg cortex, amygdala, and hypothalamus suggests that these cellular adaptations contribute to the long-term neural plastic adaptations to stress and ADTs in cortical, hypothalamic, and limbic brain structures.

Enhanced spatial discrimination learning in rats following 5,7-DHT-induced serotonergic deafferentation of the hippocampus.

Learning in rats trained in the Stone 14-unit T-maze, a complex, positively reinforced spatial discrimination task was assessed following cytotoxic (5,7-dihydroxytryptamine; 5,7-DHT) deafferentation of the serotonergic inputs to the hippocampus. Serotonergic deafferentation was accomplished by infusing the cytotoxin in to the fornix-fimbria/cingulum bundle. Lesioned rats reached criterion (i.e. learned) in significantly fewer trials and made significantly fewer errors throughout training than either vehicle-injected or sham-operated controls. This represents the first time that the effects of selective chronic destruction of serotonergic inputs to the hippocampus have been investigated. The present results provide, therefore, evidence in support of a neuromodulatory role for serotonin (5-HT) within the rat hippocampus in the mediation of the processes underlying learning and memory for this task. Other studies are, therefore, warranted in order to determine whether hippocampal 5-HT also plays a role in the mediation of the processes underlying learning and memory in other types of tasks.

Neurochemical interactions of cocaine with dopaminergic systems

Abstract The psychotropic effects of cocaine are usually attibuted to its ability to block the reuptake of dopamine into mesocortical or mesulimblic neurons, rather than to its ability to block the reuptake of 5-HT and norepinephrine. However, the interactions between cocaine and dopamine receptors have only recently been investigated. In this article , Matthew Galloway reviews new biochemical findings on the site of action of cocaine and its effects on dopamine systems. These support a role of dopamine reuptake inhibition in cocaine addiction, but the exact mechanism of interaction between the cocaine binding site and the dopamine transporter remains to be determined .

Nitric oxide and potassium chloride-facilitated striatal dopamine efflux in vivo : role of calcium-dependent release mechanisms

Previous studies investigating the calcium-dependency of nitric oxide-facilitated striatal dopamine efflux have produced conflicting results. In the current study, we have investigated the role of extracellular calcium in nitric oxide and potassium chloride-evoked striatal dopamine efflux in vivo using microdialysis. Dialysis probes were implanted in the anterior dorsal striatum of chloral hydrate-anesthetized rats. Intrastriatal infusion (20 min fraction) of the nitric oxide generators sodium nitroprusside (200 microM, 500 microM, or 1 mM) and 3-morpholinosydnonimine (1 mM) increased extracellular dopamine levels. The facilitatory effects of 3-morpholinosydnonimine and potassium chloride on dopamine efflux were attenuated following pretreatment (100 min) and co-infusion of calcium free artificial cerebral spinal fluid containing magnesium chloride. Local potassium chloride infusion (100 mM) administered alone elevated striatal dopamine efflux to a similar degree as potassium chloride (100 mM) delivered 60 min after 3-morpholinosydnonimine infusion. These results demonstrate that like potassium chloride, nitric oxide facilitates striatal dopamine efflux in vivo via a mechanism largely dependent on extracellular calcium. Also, as intrastriatal potassium chloride infusion evoked similar increases in extracellular dopamine levels in controls and subjects receiving pretreatment with the NO-generator 3-morpholinosydnonimine, it is unlikely that the functional integrity of DA nerve terminals is compromised via a neurotoxic disruption of plasma membrane potential following enhanced striatal NO production.

Progressive behavioral deficits in DJ-1-deficient mice are associated with normal nigrostriatal function.

Loss-of-function mutations in the DJ-1 gene account for an autosomal recessive form of Parkinsons disease (PD). To investigate the physiological functions of DJ-1 in vivo, we generated DJ-1 knockout (DJ-1(-/-)) mice. Younger (<1 year) DJ-1(-/-) mice were hypoactive and had mild gait abnormalities. Older DJ-1(-/-), however, showed decreased body weight and grip strength and more severe gait irregularities compared to wild-type littermates. The basal level of extracellular dopamine, evoked dopamine release and dopamine receptor D2 sensitivity appeared normal in the striatum of DJ-1(-/-) mice, which was consistent with similar results between DJ-1(-/-) and controls in behavioral paradigms specific for the dopaminergic system. An examination of spinal cord, nerve and muscle tissues failed to identify any pathological changes that were consistent with the noted motor deficits. Taken together, our findings suggest that loss of DJ-1 leads to progressive behavioral changes without significant alterations in nigrostriatal dopaminergic and spinal motor systems.

Single prolonged stress decreases glutamate, glutamine, and creatine concentrations in the rat medial prefrontal cortex

Application of single prolonged stress (SPS) in rats induces changes in neuroendocrine function and arousal that are characteristic of post traumatic stress disorder (PTSD). PTSD, in humans, is associated with decreased neural activity in the prefrontal cortex, increased neural activity in the amygdala complex, and reduced neuronal integrity in the hippocampus. However, the extent to which SPS models these aspects of PTSD has not been established. In order to address this, we used high-resolution magic angle spinning proton magnetic resonance spectroscopy (HR-MAS (1)H MRS) ex vivo to assay levels of neurochemicals critical for energy metabolism (creatine and lactate), excitatory (glutamate and glutamine) and inhibitory (gamma amino butyric acid (GABA)) neurotransmission, and neuronal integrity (N-acetylaspartate (NAA)) in the medial prefrontal cortex (mPFC), amygdala complex, and hippocampus of SPS and control rats. Glutamate, glutamine, and creatine levels were decreased in the mPFC of SPS rats when compared to controls, which suggests decreased excitatory tone in this region. SPS did not alter the neurochemical profiles of either the hippocampus or amygdala. These data suggest that SPS selectively attenuates excitatory tone, without a disruption of neuronal integrity, in the mPFC.

Glutaminase-Deficient Mice Display Hippocampal Hypoactivity, Insensitivity to Pro-Psychotic Drugs and Potentiated Latent Inhibition: Relevance to Schizophrenia

Dysregulated glutamatergic neurotransmission has been strongly implicated in the pathophysiology of schizophrenia (SCZ). Recently, presynaptic modulation of glutamate transmission has been shown to have therapeutic promise. We asked whether genetic knockdown of glutaminase (gene GLS1) to reduce glutamatergic transmission presynaptically by slowing the recycling of glutamine to glutamate, would produce a phenotype relevant to SCZ and its treatment. GLS1 heterozygous (GLS1 het) mice showed about a 50% global reduction in glutaminase activity, and a modest reduction in glutamate levels in brain regions relevant to SCZ pathophysiology, but displayed neither general behavioral abnormalities nor SCZ-associated phenotypes. Functional imaging, measuring regional cerebral blood volume, showed hippocampal hypometabolism mainly in the CA1 subregion and subiculum, the inverse of recent clinical imaging findings in prodromal and SCZ patients. GLS1 het mice were less sensitive to the behavioral stimulating effects of amphetamine, showed a reduction in amphetamine-induced striatal dopamine release and in ketamine-induced frontal cortical activation, suggesting that GLS1 het mice are resistant to the effects of these pro-psychotic challenges. Moreover, GLS1 het mice showed clozapine-like potentiation of latent inhibition, suggesting that reduction in glutaminase has antipsychotic-like properties. These observations provide further support for the pivotal role of altered glutamatergic synaptic transmission in the pathophysiology of SCZ, and suggest that presynaptic modulation of the glutamine–glutamate pathway through glutaminase inhibition may provide a new direction for the pharmacotherapy of SCZ.

Neurochemical, hormonal, and behavioral effects of chronic unpredictable stress in the rat

The high comorbidity of anxiety and depression suggests a potential degree of commonality in their etiologies. The chronic unpredictable stress (CUS) model effectively replicates depressive-like phenotypes; however, the ability of CUS to produce anxiety-like behaviors has not been adequately addressed. Using the CUS paradigm (2 stressors per day for 10 days) in adult Sprague-Dawley rats we identified behavioral, hormonal, and neurochemical changes one day after the cessation of treatment. Stress attenuated weight gain throughout the study and increased locomotor activity one day after treatment, but had no effect on anxiety-behavior as measured by the elevated plus maze. In addition, plasma corticosterone levels were positively correlated with hypothalamic serotonin (5-HT) activity one day after stress treatment as determined by the ratio of the metabolite 5-hydroxyindoleacetic acid (5-HIAA) to the parent compound (5-HIAA/5-HT ratio). These data suggest behavioral phenotypes associated with depression, but not comorbid anxiety, emerge in the immediate period after cessation of stress and that stress related physiology is related to 5-HT activity in the hypothalamus.

Methionine sulfoximine, an inhibitor of glutamine synthetase, lowers brain glutamine and glutamate in a mouse model of ALS

In an effort to alter the levels of neurochemicals involved in excitotoxicity, we treated mice with methionine sulfoximine (MSO), an inhibitor of glutamine synthetase. Since glutamate toxicity has been proposed as a mechanism for the degeneration of motor neurons in a variety of neurodegenerative diseases, we tested the effects of MSO on the transgenic mouse that overexpresses the mutant human SOD1(G93A) gene, an animal model for the primary inherited form of the human neurodegenerative disease amyotrophic lateral sclerosis (ALS). This treatment in vivo reduced glutamine synthetase activity measured in vitro by 85%. Proton magnetic resonance spectroscopy, with magic angle spinning of intact samples of brain tissue, showed that MSO treatment reduced brain levels of glutamine by 60% and of glutamate by 30% in both the motor cortex and the anterior striatum, while also affecting levels of GABA and glutathione. Kaplan-Meyer survival analysis revealed that MSO treatment significantly extended the lifespan of these mice by 8% (p<0.01). These results show that in the SOD1(G93A) model of neurodegenerative diseases, the concentration of brain glutamate (determined with (1)H-MRS) can be lowered by inhibiting in vivo the synthesis of glutamine with non-toxic doses of MSO.