Robert T. Kennedy

Dynamic Observation of Dopamine Autoreceptor Effects in Rat Striatal Slices

Abstract: Fast‐scan cyclic voltammetry has been used to measure dopamine (DA) synaptic overflow in slices of rat caudate nucleus induced by electrical stimulation with one‐, two‐, and 50‐pulse, 10‐Hz trains. Synaptic overflow in this preparation is shown to be the result of the competing effects of release and cellular uptake. Release caused by all pulses was attenuated by the D2 agonist quinpirole (1 μM). The rapid time response of the measurements (100 ms) allows the autoinhibition induced by endogenous, released DA to be resolved in real time. The concentration of DA released during the second pulse of a train was 58% of that released by the first pulse, an effect that is partially blocked by the addition of 2 μM sulpiride, a D2 antagonist, to the perfusion buffer. DA release during the first stimulus pulse is unaffected by 2 μM sulpiride, suggesting that autoreceptors are not normally occupied in this preparation. Release caused by the third pulse was 14% of the first pulse and also could be partially enhanced by 2 μM sulpiride. The duration of the inhibition of release induced by endogenous DA was estimated by varying the interval between one‐pulse stimulations until the overflow of DA induced by the second pulse was equal to that on the first; a half‐time of ∼ 17 s was found. The addition of picrotoxin (100 μM) and glutamate (10 μM) to the perfusion buffer did not affect stimulated release of DA, although the addition of atropine (100 μM) attenuated overflow for all the trains tested.

Chemical analysis of single neurons by open tubular liquid chromatography

A method, based on open tubular liquid chromatography with voltammetric detection, has been developed for the determination of trace levels of organic compounds in single cells. Individual neurons from the subesophageal ganglia ofHelix aspersa were analyzed using this method. The putative neurotransmitters, dopamine and serotonin, and their precursor amino acids, tyrosine and tryptophan, were identified and quantified in the cells.

Evoked Neuronal Activity Accompanied by Transmitter Release Increases Oxygen Concentration in Rat Striatum in vivo but Not in vitro

Dopamine and oxygen (O2) were measured in the caudate nucleus of anesthetized rats and in striatal slices during electrical stimulation. Simultaneous electrochemical detection of dopamine and O2 was accomplished with fast-scan cyclic voltammetry at a Nafion-coated carbon-fiber microelectrode. Stimulation of the medial forebrain bundle resulted in synaptic overflow of dopamine in the caudate nucleus. At the same time, O2 concentration increased in the extracellular fluid with two separate phases. The amplitude of the initial increase directly correlated with the frequency of the stimulus, with the time of maximum concentration reproducible across a range of frequencies. The second increase occurred at later times with a more random amplitude and with a broad, variable shape. Agents which blocked vasodilation affected both phases: Atropine attenuated the initial increase, while the second feature was nearly absent after theophylline. Yohimbine and α-methyl-p-tyrosine did not affect the O2 responses. Local electrical stimulation of the slice preparation also resulted in dopamine overflow, but a prolonged decrease in O2 concentration accompanied this event. Striatal field stimulation in vivo produced changes in O2 concentration dependent on the relative position of the stimulating and working electrodes, but none of the responses resembled that seen in the caudate slice. Thus, while measurements in brain slices show O2 consumption as a result of stimulated neuronal activity, an apparent elevation of local cerebral blood flow during and after stimulation dominate the in vivo response.

Microelectrodes in Biological Systems

The small size of microelectrodes enables their use as sensors in biological systems of easily oxidized chemical substances. Carbon-fiber electrodes implanted in the brain can be used to examine the dynamic concentration changes of the neurotransmitter dopamine. In thin slices of brain tissue, the electrodes can be used to simultaneously detect neurotransmitter secretion and oxygen use. At single cells secretion processes can be resolve in real time.