r3,-Adrenergic Regulation of Cyclic Nucleotide Levels and Potassium Fluxes in Rat Parotid Gland

It is generally considered that both w and i3-adrenoceptors exist in rat parotid glands (1, 2). In our previous paper, the effect of adrenergic stimuli on spontaneous potassium fluxes was found to be distinctly different between at and i3-adrenergic agonists, the former increases potassium efflux and the latter decreases potassium release (3, 4). The present in vitro investigation was undertaken in an attempt to obtain further information as to the role of i9-adrenoceptor subtypes on isoproterenol-induced decrease in potassium release from rat parotid tissue. Male Wistar rats (260-280 g) were used throughout the experiments. Parotid glands were removed under pentobarbital (40 mg/ kg, i.p.) anesthesia, and then the parotid slices were prepared and incubated according to the procedures described by Ohshika et al. (5). Briefly, the slices were incubated for 30 min at 37°C in the Krebs-Ringer bicarbonate (KRB) medium gassed with a 95% 02-5% C02 mixture (equilibration period). Then, they were incubated for 5 min with isopro terenol or selective ;3-adrenergic agonists (challenge). i3-Adrenergic antagonists were added into the incubation medium 2 min before the addition of isoproterenol (pretreat ment). At the end of the final incubation, 100 /€l aliquots of the medium were removed, and the slices were homogenized together with the remaining medium. The homogenates were centrifuged at 1000 x g for 10 min. The potassium concentrations in aliquots of the medium and the supernatants were deter mined by a flame photometer (Corning) using a lithium internal standard. The release of potassium was expressed as the percent of total potassium in the slices, using the formula described by Martinez et al. (6). The cyclic AMP content of parotid tissue was measured after a 5 min incubation of the parotid slices in KRB medium with various h3-adrenergic agonists. After the incubation, the tissue slices were immediately frozen with liquid nitrogen and stored until time for measurement. Cyclic AMP was assayed by radioimmunoassay (7) with a cyclic AMP assay kit (Yamasa Shoyu Co., Japan). Data are presented as the mean+S .E. Statistical analysis were performed using Student's t-test (two-tailed). The following drugs were used: (-) isoproterenol HCI (Sigma), (+)-propranolol HCI (Sigma), and butoxamine HCI (Bur roughs-Wellcome). Dobutamine HCI was a gift from the Shionogi Pharmaceutical Co. (Japan), procaterol HCI was a gift from Otsuka Pharmaceutical Co. (Japan), and metoprolol HCI was a gift from Fujisawa Pharmaceutical Co. (Japan). Table 1 shows the effects of various ~~ adrenoceptor agents on …