Effects of clonidine on the stimulation-evoked release of3H-noradrenaline from superfused rat brain slices as a function of the biophase concentration

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Abstract
The influence of clonidine on the stimulation-evoked overflow of tritium was studied in brain slices preincubated with3H-noradrenaline. The slices were prepared from parietal cortex (Cx), nucleus anterior hypothalami (nah) and nucleus tractus solitarii (nts). After preincubation, the tissues were superfused at 23°C or 37°C with a medium containing the noradrenaline uptake inhibitor desipramine. Electrical field stimulation was applied using stimulation frequencies of 0.3–10 Hz. At 23°C/0.3 Hz, clonidine concentration-dependently inhibited the evoked overflow of tritium in all three brain regions. In contrast, at 23°C/3 Hz the inhibitory effect of the drug in the Cx was abolished and a facilitation was observed in the nah and nts. When tested at increasing frequencies of stimulation in the nts at 23°C, clonidine exerted a dual action, characterized by a reduction of electrically evoked responses at frequencies below 1 Hz and a facilitation at frequencies above 1 Hz. At 37°C, clonidine concentration-dependently decreased the evoked overflow in all brain regions studied, this effect being more pronounced at 0.3 Hz than at 3 Hz. The apparent lack of an effect of clonidine on the stimulation-evoked overflow of tritium in the Cx at 23°C/3 Hz was turned to a facilitation when noradrenaline (0.01 μmol/l) was included in the superfusion medium. Conversely, an inhibitory effect of clonidine was seen when the uptake blocker desipramine (as well as noradrenaline) was omitted from the superfusion medium. The facilitation by clonidine of the stimulus-induced release in the nah at 23°C/3 Hz was reversed to inhibition when the noradrenaline content of the slices was reduced to about 60% by exposure to a medium containing 50 mmol/l potassium before preincubation with3H-noradrenaline. A marked difference regarding the two superfusion temperatures was noted at the ultrastructural level: the superfusion of slices resulted in a widening of extracellular spaces, dissociation of synaptic clefts with preservation of presynaptic terminals, and intracytoplasmatic uptake of horseradish peroxidase. These changes were more pronounced after superfusion at 37°C than at 23°C. In accordance with these observations, the retention of3H-polyethyleneglycol in slices superfused at 37°C was greater than in slices superfused at 23°C. It is concluded that in superfusion/stimulation experiments with brain slices temperature-dependent changes at the ultrastructural level may have an influence on the concentration of noradrenaline in the synaptic cleft. The results emphasize the role of the biophase concentration of noradrenaline with regard to the magnitude and the direction of the presynaptic effect of clonidine.

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