Paul A. Garris

Dissociation of dopamine release in the nucleus accumbens from intracranial self-stimulation

Mesolimbic dopamine-releasing neurons appear to be important in the brain reward system,. One behavioural paradigm that supports this hypothesis is intracranial self-stimulation (ICS), during which animals repeatedly press a lever to stimulate their own dopamine-releasing neurons electrically. Here we study dopamine release from dopamine terminals in the nucleus accumbens core and shell in the brain by using rapid-responding voltammetric microsensors during electrical stimulation of dopamine cell bodies in the ventral tegmental area/substantia nigra brain regions. In rats in which stimulating electrode placement failed to elicit dopamine release in the nucleus accumbens, ICS behaviour was not learned. In contrast, ICS was acquired when stimulus trains evoked extracellular dopamine in either the core or the shell of the nucleus accumbens. In animals that could learn ICS, experimenter-delivered stimulation always elicited dopamine release. In contrast, extracellular dopamine was rarely observed during ICS itself. Thus, although activation of mesolimbic dopamine-releasing neurons seems to be a necessary condition for ICS, evoked dopamine release is actually diminished during ICS. Dopamine may therefore be a neural substrate for novelty or reward expectation rather than reward itself.

Real-time decoding of dopamine concentration changes in the caudate-putamen during tonic and phasic firing

The fundamental process that underlies volume transmission in the brain is the extracellular diffusion of neurotransmitters from release sites to distal target cells. Dopaminergic neurons display a range of activity states, from low-frequency tonic firing to bursts of high-frequency action potentials (phasic firing). However, it is not clear how this activity affects volume transmission on a subsecond time scale. To evaluate this, we developed a finite-difference model that predicts the lifetime and diffusion of dopamine in brain tissue. We first used this model to decode in vivo amperometric measurements of electrically evoked dopamine, and obtained rate constants for release and uptake as well as the extent of diffusion. Accurate predictions were made under a variety of conditions including different regions, different stimulation parameters and with uptake inhibited. Second, we used the decoded rate constants to predict how heterogeneity of dopamine release and uptake sites would affect dopamine concentration fluctuations during different activity states in the absence of an electrode. These simulations show that synchronous phasic firing can produce spatially and temporally heterogeneous concentration profiles whereas asynchronous tonic firing elicits uniform, steady-state dopamine concentrations.

Real-time measurement of electrically evoked extracellular dopamine in the striatum of freely moving rats

Abstract: The real‐time measurement of electrically evoked dopamine was established in brain extracellular fluid of freely moving rats. Dopamine was monitored by fast‐scan cyclic voltammetry at carbon fiber microelectrodes lowered into the striatum by means of a detachable micromanipulator. A stimulating electrode, previously implanted in the substantia nigra, was used to evoke striatal dopamine efflux. Evoked extracellular dopamine was both current and frequency dependent. When low current intensities (±125 µA) and frequencies (10–20 Hz) were applied, detectable levels of dopamine were elicited without a perceptible behavioral response. Reproducible concentrations of extracellular dopamine could be evoked in the same rat for at least 2 months. These concentrations, moreover, were significantly higher in freely moving rats compared with rats anesthetized with Equithesin. Analysis of measured curves for dopamine uptake and release rates revealed that anesthesia inhibits release but does not affect uptake. It is concluded that (a) fast‐scan cyclic voltammetry at carbon fiber microelectrodes is a viable technique for the measurement of electrically evoked dopamine in brain extracellular fluid of freely moving rats, (b) it is possible to determine in situ rate constants for dopamine release and uptake from these temporally and spatially resolved measurements of levels of dopamine, and (c) transient changes in extracellular dopamine levels elicited by electrical stimulation are affected by anesthesia.

Determination of release and uptake parameters from electrically evoked dopamine dynamics measured by real-time voltammetry

Quantifying mechanisms underlying extracellular signaling by the neurotransmitter dopamine (DA) is a difficult task, particularly in the complex extracellular microenvironment of the intact brain. In this study, two methods for evaluating release and uptake from DA dynamics monitored by real-time voltammetry are described. Both are based on a neurochemical model characterizing electrically evoked levels of DA as a balance between these opposing mechanisms. The theoretical basis of what is called here nonlinear regression and single curve analyses is given. Fitting simulated data tests the reliability of the methods. The two analyses are also compared with an experimental data set describing the effects of pharmacologically inhibiting the DA transporter in the caudate-putamen (CP) and nucleus accumbens (NAc). The results indicate that nonlinear regression and single curve analyses are suitable for quantifying release and uptake mechanisms underlying DA neurotransmission. Additionally, the most important experimental finding of this technical study was the independent confirmation of high affinity (approximately 0.2 microM) DA uptake in the intact striatum.

Real-time decoding of dopamine concentration changes in the caudate-putamen during tonic and phasic firing: Decoding dopamine neurotransmission

The fundamental process that underlies volume transmission in the brain is the extracellular diffusion of neurotransmitters from release sites to distal target cells. Dopaminergic neurons display a range of activity states, from low‐frequency tonic firing to bursts of high‐frequency action potentials (phasic firing). However, it is not clear how this activity affects volume transmission on a subsecond time scale. To evaluate this, we developed a finite‐difference model that predicts the lifetime and diffusion of dopamine in brain tissue. We first used this model to decode in vivo amperometric measurements of electrically evoked dopamine, and obtained rate constants for release and uptake as well as the extent of diffusion. Accurate predictions were made under a variety of conditions including different regions, different stimulation parameters and with uptake inhibited. Second, we used the decoded rate constants to predict how heterogeneity of dopamine release and uptake sites would affect dopamine concentration fluctuations during different activity states in the absence of an electrode. These simulations show that synchronous phasic firing can produce spatially and temporally heterogeneous concentration profiles whereas asynchronous tonic firing elicits uniform, steady‐state dopamine concentrations.

Microdialysis of dopamine interpreted with quantitative model incorporating probe implantation trauma

Although microdialysis is widely used to sample endogenous and exogenous substances in vivo, interpretation of the results obtained by this technique remains controversial. The goal of the present study was to examine recent criticism of microdialysis in the specific case of dopamine (DA) measurements in the brain extracellular microenvironment. The apparent steady‐state basal extracellular concentration and extraction fraction of DA were determined in anesthetized rat striatum by the concentration difference (no‐net‐flux) microdialysis technique. A rate constant for extracellular clearance of DA calculated from the extraction fraction was smaller than the previously determined estimate by fast‐scan cyclic voltammetry for cellular uptake of DA. Because the relatively small size of the voltammetric microsensor produces little tissue damage, the discrepancy between the uptake rate constants may be a consequence of trauma from microdialysis probe implantation. The trauma layer has previously been identified by histology and proposed to distort measurements of extracellular DA levels by the no‐net‐flux method. To address this issue, an existing quantitative mathematical model for microdialysis was modified to incorporate a traumatized tissue layer interposed between the probe and surrounding normal tissue. The tissue layers are hypothesized to differ in their rates of neurotransmitter release and uptake. A post‐implantation traumatized layer with reduced uptake and no release can reconcile the discrepancy between DA uptake measured by microdialysis and voltammetry. The model predicts that this trauma layer would cause the DA extraction fraction obtained from microdialysis in vivo calibration techniques, such as no‐net‐flux, to differ from the DA relative recovery and lead to an underestimation of the DA extracellular concentration in the surrounding normal tissue.

Sub-second changes in accumbal dopamine during sexual behavior in male rats

Transient (200–900 ms), high concentrations (200–500 nM) of dopamine, measured using fast-scan cyclic voltammetry, occurred in the nucleus accumbens core of male rats at the presentation of a receptive female. Additional dopamine signals were observed during subsequent approach behavior. Background-subtracted cyclic voltammograms of the naturally-evoked signals matched those of electrically-evoked dopamine measured at the same recording sites. Administration of nomifensine amplified natural and evoked dopamine release, and increased the frequency of detectable signals. While gradual changes in dopamine concentration during sexual behavior have been well established, these findings dramatically improve the time resolution. The observed dopamine transients, probably resulting from neuronal burst firing, represent the first direct correlation of dopamine with sexual behavior on a sub-second time scale.

Dopamine release and uptake rates both decrease in the partially denervated striatum in proportion to the loss of dopamine terminals

The present study tested the hypothesis that normal concentrations of extracellular dopamine are preserved in the partially denervated striatum without active compensatory changes in dopamine uptake or release. One to four weeks after adult rats were unilaterally lesioned with 6-hydroxydopamine, fast-scan cyclic voltammetry at Nafion-coated, carbon-fiber microelectrodes was used to monitor extracellular dopamine levels in vivo, under urethane anesthesia. Simultaneous voltammetric recordings were collected in the lesioned and contralateral control striata. Extracellular dopamine was elicited by bilateral electrical stimulation of the medial forebrain bundle. A 20 Hz stimulation evoked similar concentrations of extracellular dopamine in both lesioned and control striata, although tissue dopamine was decreased 30-70% in lesioned striata, as determined subsequently by HPLC-EC. However, kinetic analysis of the voltammetric recordings revealed that the concentration of dopamine released per stimulus pulse and Vmax for dopamine uptake decreased in proportion to the magnitude of the lesion. These data support the hypothesis that normal extracellular dopamine levels can be generated in the partially lesioned striatum in the absence of active neuronal compensation. These results also suggest that passive mechanisms involved in the regulation of extracellular dopamine play an important role in maintaining function during the preclinical or presymptomatic phase of Parkinsons disease.

A ROLE FOR PRESYNAPTIC MECHANISMS IN THE ACTIONS OF NOMIFENSINE AND HALOPERIDOL

Psychomotor stimulants and neuroleptics exert multiple effects on dopaminergic signaling and produce the dopamine (DA)-related behaviors of motor activation and catalepsy, respectively. However, a clear relationship between dopaminergic activity and behavior has been very difficult to demonstrate in the awake animal, thus challenging existing notions about the mechanism of these drugs. The present study examined whether the drug-induced behaviors are linked to a presynaptic site of action, the DA transporter (DAT) for psychomotor stimulants and the DA autoreceptor for neuroleptics. Doses of nomifensine (7 mg/kg i.p.), a DA uptake inhibitor, and haloperidol (0.5 mg/kg i.p.), a dopaminergic antagonist, were selected to examine characteristic behavioral patterns for each drug: stimulant-induced motor activation in the case of nomifensine and neuroleptic-induced catalepsy in the case of haloperidol. Presynaptic mechanisms were quantified in situ from extracellular DA dynamics evoked by electrical stimulation and recorded by voltammetry in the freely moving animal. In the first experiment, the maximal concentration of electrically evoked DA ([DA](max)) measured in the caudate-putamen was found to reflect the local, instantaneous change in presynaptic DAT or DA autoreceptor activity according to the ascribed action of the drug injected. A positive temporal association was found between [DA](max) and motor activation following nomifensine (r=0.99) and a negative correlation was found between [DA](max) and catalepsy following haloperidol (r=-0.96) in the second experiment. Taken together, the results suggest that a dopaminergic presynaptic site is a target of systemically applied psychomotor stimulants and regulates the postsynaptic action of neuroleptics during behavior. This finding was made possible by a voltammetric microprobe with millisecond temporal resolution and its use in the awake animal to assess release and uptake, two key mechanisms of dopaminergic neurotransmission. Moreover, the results indicate that presynaptic mechanisms may play a more important role in DA-behavior relationships than is currently thought.

Amphetamine Paradoxically Augments Exocytotic Dopamine Release and Phasic Dopamine Signals

Drugs of abuse hijack brain-reward circuitry during the addiction process by augmenting action potential-dependent phasic dopamine release events associated with learning and goal-directed behavior. One prominent exception to this notion would appear to be amphetamine (AMPH) and related analogs, which are proposed instead to disrupt normal patterns of dopamine neurotransmission by depleting vesicular stores and promoting nonexocytotic dopamine efflux via reverse transport. This mechanism of AMPH action, though, is inconsistent with its therapeutic effects and addictive properties, which are thought to be reliant on phasic dopamine signaling. Here we used fast-scan cyclic voltammetry in freely moving rats to interrogate principal neurochemical responses to AMPH in the striatum and relate these changes to behavior. First, we showed that AMPH dose-dependently enhanced evoked dopamine responses to phasic-like current pulse trains for up to 2 h. Modeling the data revealed that AMPH inhibited dopamine uptake but also unexpectedly potentiated vesicular dopamine release. Second, we found that AMPH increased the amplitude, duration, and frequency of spontaneous dopamine transients, the naturally occurring, nonelectrically evoked, phasic increases in extracellular dopamine. Finally, using an operant sugar reward paradigm, we showed that low-dose AMPH augmented dopamine transients elicited by sugar-predictive cues. However, operant behavior failed at high-dose AMPH, which was due to phasic dopamine hyperactivity and the decoupling of dopamine transients from the reward predictive cue. These findings identify upregulation of exocytotic dopamine release as a key AMPH action in behaving animals and support a unified mechanism of abused drugs to activate phasic dopamine signaling.