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The ability of lanthanum (La3+) to block calcium efflux from smooth muscle cells of the rabbit aorta has been examined. La3+ promotes the very early phase of 45Ca efflux, which is extracellular in origin, and partially inhibits the latter, cellular portion. Stimulation of 45Ca efflux caused by the release of intracellular 45Ca with either 10(-4) M dinitrophenol or 10 mM caffeine was not reduced by pretreatment with 10 mM La for 40 min, whereas stimulation due to norepinephrine was abolished. It was concluded that during the use of the "La method" for measuring cellular 45Ca there is an underestimation due to unblocked 45Ca loss. This loss can be reduced by processing tissue at 2 degrees C, which inhibits transport processes. The time course of 45Ca uptake and the stimulation of uptake by high K+ are qualitatively but not quantitatively similar if tissues are washed at 37 and 2 degrees C. Tissues washed in La3+ at 2 degrees C for 60 min retain approximately double the cellular 45Ca of those washed at 37 degrees C. This methodology provides an improved correlation between estimates of cellular calcium derived from 45Ca uptake and 45Ca efflux experiments.

The ability of norepinephrine (NE), caffeine (CAF), theophylline (THEO), dinitrophenol (DNP), and potassium (high K) to mobilize cellular Ca2+ in rabbit aorta was examined using 45Ca-efflux techniques. After 10 min of Ca2+ deprivation using either Ca-free EGTA or Ca-free lanthanum (La3+) buffers, NE, CAF, and DNP still caused an increase in 45Ca-efflux rate, indicating a cellular source of 45Ca, while high K did not. Contractile behavior after Ca removal paralleled 45Ca-efflux events. CAF (10 mM) inhibited NE contractile responses, and this inhibition was associated with the depletion of the NE-releasable Ca2+ store. Previous exposure to CAF during 45Ca efflux reduced subsequent stimulation of 45Ca efflux was not additive. THEO caused a stimulation of 45Ca efflux similar to CAF. CAF, THEO, and 3-isobutyl-l-methylxanthine caused a two- to threefold increase in cAMP levels in association with their stimulation of 45Ca efflux. These results suggest that NE and methylxanthines mobilize a common cellular Ca2+ source that is associated with contraction in the case of NE but relaxation in the case of methylxanthines.

A sustained increase in sympathetic nervous system (SNS) activity was induced by substituting a 10% sucrose solution for the drinking water of rats fed laboratory chow ad libitum. The effects of increased SNS activity on alpha 1-adrenergic processes in liver were examined by evaluating three alpha 1-responses, namely, phenylephrine-stimulated ouabain-sensitive 86Rb+ uptake, 45Ca2+ efflux, and glucose release. Sucrose feeding abolished phenylephrine stimulation of ouabain-sensitive 86Rb+ uptake and 45Ca2+ efflux and induced a three- to fourfold reduction in the ability of phenylephrine to stimulate glucose release from liver slices. Pretreatment with 6-hydroxydopamine markedly reduced liver norepinephrine content. When 6-hydroxydopamine was used to prevent the sucrose-induced increase in SNS activity, the changes in 86Rb+ uptake, 45Ca2+ efflux, and glucose release that otherwise followed sucrose feeding were not observed. Sucrose feeding did not alter binding of the alpha 1-antagonist [3H]prazosin to liver cell membrane alpha 1-receptors or displacement of [3H]prazosin by the alpha-agonist epinephrine. These observations suggest that sustained increases in SNS activity may have profound effects on liver alpha 1-adrenergic events that occur subsequent to hormone-receptor interaction.

To evaluate the influence of intracellular alkalinization on basal and agonist-induced vascular tone, we studied the effect of NH4Cl on rat aorta. NH4Cl induced a gradually developing contraction in a dose-dependent manner. Although the contractile response to 20 mM NH4Cl was associated with a latent period (LP) of 23.4 +/- 2.8 min, intracellular pH (pHi) measurements in cultured rat aortic smooth muscle cells showed that NH4Cl-induced intracellular alkalinization was immediate and transient, returning to basal pHi levels in about 30-35 min. Agents that elevate Ca2+, such as A23187 and high KCl, significantly reduced the LP associated with 20 mM NH4Cl-induced contraction. NH4Cl-induced contractions were sensitive to extracellular Ca2+ removal and to the addition of forskolin (1 microM); however, NH4Cl by itself did not cause Ca2+-influx as shown by 45Ca-uptake studies. Addition of 20 mM NH4Cl to precontracted tissues resulted in a transient relaxation, which was complete in approximately 10 min, followed by a contraction above the original level of tone. NH4Cl pretreatment caused time-dependent alterations in both the rapid and slow phases of phenylephrine and angiotensin II contractions. Rapid-phase of phenylephrine and angiotensin II contractions. Rapid-phase responses were diminished at shorter NH4Cl incubation times (10 min), whereas slow-phase response was augmented after a longer incubation (20 min). Overall, the vasorelaxant and vasoconstrictor effects induced by NH4Cl suggest a complex relationship between intracellular alkalinization and arterial contractility.