Charles W. Bradberry

Simultaneous voltammetric and chemical monitoring of dopamine release in situ

The validity of voltammetric monitoring of stimulated dopamine (DA) release was tested by simultaneous chemical assay of the extracellular fluid region next to the tip of a selective Nafion-coated graphite voltammetric electrode. Micro alumina probes were positioned next to the recording electrode tip to adsorb released catecholamines. The catecholamines were desorbed into microvolumes of acid and analyzed by very sensitive HPLC assays to provide unequivocal chemical verification of the electrochemical signals. The results fully confirm that selective voltammetric electrodes can detect released DA and that the concentrations determined voltammetrically or chemically agree well.

Research reportRegulation of excitatory amino acid release by N-methyl-d-aspartate receptors in rat striatum: in vivo microdialysis studies

The microdialysis technique was utilized to study the effects of N-methyl-D-aspartate (NMDA) receptor ligands on the in vivo release of endogenous glutamate (Glu) and aspartate (Asp) from the rat striatum. Addition of NMDA (250 and 500 microM) to the dialysis perfusion solution resulted in a striking dose-dependent increase in extracellular concentrations of Glu and Asp in the striatum. The NMDA-induced effects were reduced in a dose-related way by prior perfusion with 75 microM dizocilpine (MK-801), a non-competitive NMDA receptor antagonist. MK-801, at 75 microM, produced no changes on basal levels of Glu and Asp. However, 100 microM MK-801 did increase Glu and Asp extracellular concentrations. Local infusion with 500 microM D-serine, an agonist at the glycine site associated to the NMDA receptor, significantly increased basal level of Glu, but not Asp. Such D-serine-induced effects were reduced by 7-Cl-kynurenic acid (200 microM), a selective blocker of the glycine site present in the NMDA receptor. It is proposed that activation of NMDA receptors by endogenous Glu and Asp enhances the subsequent release of these excitatory amino acids in the striatum. Part of these NMDA receptors might be located presynaptically on cortico-striatal nerve endings. In addition, postsynaptic NMDA receptors present in the striatum may also indirectly modulate the release of Glu and Asp, through trans-synaptic mechanism.

Alcohol plus Cocaine: The Whole Is More Than the Sum of Its Parts

Cocaethylene, an active metabolite that arises through hepatic transesterification of cocaine when cocaine and ethanol are used together, shares many neurochemical and pharmacological properties with cocaine. Cocaethylene is similar to cocaine in its properties as an indirect dopamine agonist, and human subjects cannot distinguish its effects from those of cocaine. Cocaethylene, and especially its isopropyl analog, are more selective indirect dopamine agonists than cocaine, with relatively weak potency at the serotonin transporter. Cocaethylene may contribute to the manifestations and consequences of combined cocaine and ethanol use, although its relative importance remains unclear.

Phasic alterations in dopamine and serotonin release in striatum and prefrontal cortex in response to cocaine predictive cues in behaving rhesus macaques.

The ability of environmental cues associated with cocaine availability to cause relapse may result from conditioned activation of dopamine (DA) release. We examined this hypothesis in macaque monkeys by conducting microdialysis studies in animals during exposure to a cocaine predictive compound cue. In addition to studying DA release in mesolimbic and sensorimotor striatum, both DA and serotonin levels were determined in the prefrontal cortex (medial orbitofrontal and anterior cingulate). The compound cue employed visual, auditory, and olfactory components, and was salient to the animals as demonstrated by anticipatory lever pressing in the absence of cocaine. During a 10-min period of exposure prior to cocaine availability, there was no significant increase in striatal or cortical DA. The addition of a DA uptake inhibitor to the striatal perfusate to reduce the potential interference of neuronal uptake did not alter the results. In contrast to the lack of any change in striatal DA, a significant decrease in extracellular serotonin in the prefrontal cortex during the 10 min of cue exposure was observed.

Orbitofrontal and Anterior Cingulate Cortex Neurons Selectively Process Cocaine-Associated Environmental Cues in the Rhesus Monkey

Encounters with stimuli associated with drug use are believed to contribute to relapse. To probe the neurobiology of environmentally triggered drug use, we have conducted single-unit recordings in rhesus monkeys during presentation of two distinct types of drug paired cues that differentially support drug-seeking. The animals were highly conditioned to these cues via exposure during self-administration procedures conducted over a 4 year period. The cues studied were a discriminative cue that signaled response-contingent availability of cocaine, and a discrete cue that was temporally paired with the cocaine infusion (0.1 or 0.5 mg/kg). Two cortical regions consistently activated by cocaine-associated cues in human imaging studies are the orbitofrontal (OFC) and anterior cingulate cortex (ACC), though little is known about cortical neuronal activity responses to drug cues. We simultaneously recorded single-unit activity in OFC and ACC as well as in dorsal striatum in rhesus monkeys during cocaine self-administration. Dorsal striatal neurons were less engaged by drug cues than cortical regions. Between OFC and ACC, distinct functionality was apparent in neuronal responses. OFC neurons preferentially responded to the discriminative cue, consistent with a role in cue-induced drug-seeking. In contrast, the ACC did not respond more to the discriminative cue than to the discrete cue. Also distinct from the OFC, ACC showed sustained firing throughout the 18 s duration of the discrete cue. This pattern of sustained activation in ACC is consistent with a role in reward expectation and/or in mediating behavioral effects of discrete cues paired with drug infusions.

α2-receptor control over release of noradrenaline in rat thalamus

Rat brain thalamic tissue was examined for the presence of alpha 2-receptor control over noradrenaline (NA) utilization. Systemically administered clonidine (0.1 mg/kg i.p.) decreased NA turnover, yohimbine (10 mg/kg i.p.) increased NA turnover, and prazosin (0.5 mg/kg i.p.) had no effect. In vitro, yohimbine increased K+-stimulated overflow of endogenous NA in a dose-dependent fashion. It appears that locus coeruleus neurons which project to thalamus exhibit the same type of alpha 2-receptor control as those terminating in other forebrain regions.

Immobilization of ascorbic acid oxidase

Ascorbic acid has long been recognized as an important vitamin [1], and has been shown to exist in high concentrations in the central nervous system of small animals and man. The regional distribution of AA in rat [2] and human [3] brain has been reported. There appear to be homeostatic mechanisms for controlling AA levels, as well as an active uptake mechanism [4]. This, in addition to the many newly reported effects of AA on neuronal tissue, of which just a few are listed, suggests an important role(s) for this molecule in the functioning of mammalian brain [5-7]. Ascorbic acid oxidase (EC 1.10.3.3, AAO) has become a useful tool in the manipulation of [AA] in neuronal tissue preparations [8,9] for the purpose of gaining information on AA directly, as well as removing it as an interference in the electrochemical determination of catecholamine neurotransmitters, serotonin, and their key metabolites. With the intention of obtaining a form of this extremely useful enzyme suitable for use in flowing stream analyses (for example, direct electrochemical analysis of neuronal tissue perfusates), we have immobilized AAO onto Sepharose 4B activated by cyanogen bromide (CNBr). In [10] the enzyme was immobilized in an enzyme electrode configuration; however, this method is unsuitable for the type of flowing stream applications inten-