Using microdialysis coupled on-line to capillary electrophoresis to study the effects of estradiol and psychostimulants on neurotransmitter release.

Abstract

One way to study brain function is to correlate changes in neurotransmitter levels with behavior and pharmacological manipulation. In such measurements temporal resolution is important to achieving good correlations because neurochemical concentrations fluctuate rapidly. Previous work coupled microdialysis on-line to capillary electrophoresis with laser induced fluorescence (CE-LIF) and demonstrated that this method can measure changes in neuroactive amine neurotransmitters (glutamate, aspartate, GABA, taurine, serine, glutamine, and glycine) every 13 seconds, realizing a 60-fold improvement in temporal resolution over typical HPLC measurements of dialysate. This improvement is due to the high mass sensitivity and automation of CE. With the CE-LIF method temporal resolution becomes limited by Taylor dispersion rather than sample collection/separation time. In this thesis, experiments investigating the mechanisms behind dopamine release regulation in the striatum demonstrate how the high temporal resolution, high sensitivity, and high throughput capabilities of the CE-LIF method can be utilized routinely in mechanistic <italic>in vivo</italic> and <italic>in vitro</italic> experiments. Estradiol has a profound effect on drug taking behavior and potentiates dopamine release in the striatum. Previous work showed that estradiol decreases stimulated GABA release in the striatum supporting the hypothesis that estradiol enhances dopamine function, therefore reinforcing addictive substances such as cocaine, by inhibiting GABA release, which in turn disinhibits dopamine release. Current work using the CE-LIF instrument expanded this mechanism by showing that both estrogen receptor-alpha (ERalpha) expression and metabotropic glutamate receptor 5 (mGluR5) activity influence the modulatory effect of estradiol on K<super>+</super> evoked GABA release. A similar disinhibition mechanism was observed <italic>in vitro</italic> between delta opiates and dopamine. SNC80, a delta opiate receptor agonist, enhances amphetamine induced efflux of dopamine from striatal tissue. Utilizing the high throughput capacity along with the multianalyte detection of the CE instrumentation we observed that SNC80 enhances glutamate and glycine efflux while it decreases GABA efflux. These results along with results from dopamine efflux assays suggest that SNC80 enhances stimulated dopamine efflux by enhancing glutamatergic activity at NMDA receptors through disinhibition of cortical glutamate terminals. These <italic>in vivo</italic> and <italic> in vitro</italic> experiments highlight the role of GABA as an important modulator of dopamine release in the striatum.