Chronic low back pain assessment using surface electromyography.

This investigation examined surface electromyography as an additional tool in the comprehensive clinical evaluation of patients with chronic low back pain (CLBP). Electromyographic signals from electrodes placed in the lumbar area of 30 CLBP patients and 30 non-pain control subjects were compared. Patients and controls were matched for age, gender, and body mass index. Paired t test showed a statistically significant difference between the two groups. The muscle activity mean values were threefold higher in CLBP patients than in controls (P < 0.00001) in the static testing, and twofold higher in CLBP patients than in controls (P < 0.00001) in the dynamic testing. Our findings indicate that surface electromyography assessment of the paraspinal muscle activity may be a useful objective diagnostic tool in the comprehensive evaluation of CLBP.

Angiotensin II receptor-mediated calcium influx in bovine adrenal glomerulosa cells.

The cytoplasmic calcium ([Ca2+]i) response to angiotensin II (AII) in bovine adrenal glomerulosa cells is characterized by an initial transient peak due to intracellular Ca2+ mobilization, followed by a sustained plateau phase that is dependent on Ca2+ entry from the extracellular fluid. In Fura-2 loaded cells, the calcium channel antagonists, nifedipine (1 microM) and verapamil (20 microM), only partially reduced the cytosolic calcium profile induced by AII. The dihydropyridine agonist, Bay K 8644, caused a moderate increase in [Ca2+]i when added in concentrations of 50-100 nM, but did not enhance the AII-induced rise in [Ca2+]i. These results indicate that most of the AII-stimulated Ca2+ influx is through channels that are insensitive to dihydropyridines and verapamil. In contrast, the inorganic Ca2+ channel blocker, LaCl3 (10 microM), inhibited the AII-induced plateau phase by more than 50%. The AII-induced Ca2+ signal was not affected by treatment with pertussis toxin (100-300 ng/ml for 12 h). The prior addition of specific AII-antagonists (DuP 753, a nonpeptide antagonist, and three peptide analogs, [Sar1,Thr8]AII, [Sar1,Ala8]AII, and [Sar1,Ile8]AII) completely inhibited the AII-induced Ca2+ signal. However, addition of up to 25,000 molar excess of these antagonists at intervals from 10 sec to 5 min after AII caused progressively less attenuation of the sustained Ca2+ signal. After 5 min, addition of antagonists did not inhibit the agonist-induced Ca2+ response for up to 20 min. The progressive loss of ability of the antagonists to inhibit the sustained elevation of [Ca2+]i could reflect prolonged activation of the receptor or of a subsequent process that maintains Ca2+ influx despite receptor blockade. It is possible that sequestration and/or endocytosis of the AII-receptor complex is accompanied by continued generation of intracellular signals that contribute to the maintenance of the [Ca2+]i response.

The mas oncogene enhances angiotensin-induced [Ca2+]i responses in cells with pre-existing angiotensin II receptors.

The proposal that the mas oncogene is an angiotensin receptor was evaluated in Xenopus oocytes injected with human and rat mas RNA transcripts, and during transient expression of mas in several cell lines. No evidence of mas-induced angiotensin II (AII) receptors or [Ca2+]i responses was observed in Xenopus oocytes or in most of the transfected cells. However, Cos-1 cells, which showed a small endogenous [Ca2+]i response to AII, exhibited a modest but reproducible enhancement of this response after mas transfection. Such responses were inhibited by [Sar1, Ala8]AII and [Sar1, Ile8]AII, but not by [D-Arg1, D-Pro2, D-Trp7,9, Leu11] substance P, an antagonist reported to inhibit mas-induced responses to AII in oocytes. These findings are not compatible with the proposal that the mas oncogene is an angiotensin receptor, but suggest that expression of mas leads to increased responsiveness of the endogenous AII signaling system.

Relationship between agonist- and thapsigargin-sensitive calcium pools in adrenal glomerulosa cells. Thapsigargin-induced Ca2+ mobilization and entry.

The relationships between agonist-sensitive calcium pools and those discharged by the Ca(2+)-ATPase inhibitor thapsigargin were studied in intact bovine adrenal glomerulosa cells and a subcellular adrenocortical membrane fraction. In Fura-2-loaded glomerulosa cells, angiotensin II (AII) stimulated a rapid increase in cytoplasmic Ca2+ concentration ([Ca2+]i) followed by a smaller plateau phase that was dependent on extra-cellular Ca2+. In such cells thapsigargin caused a sustained and dose-dependent increase in [Ca2+]i which was diminished in Ca(2+)-deficient medium. The contribution of an influx component to the thapsigargin-induced [Ca2+]i response was demonstrated by measurement of 45Ca influx rate in glomerulosa cells. Thapsigargin-induced Ca2+ entry was significantly less than that evoked by AII, and its kinetics were similar to those of the concomitant increase in [Ca2+]i. The rate of emptying of the agonist-responsive Ca2+ pool after thapsigargin treatment, as indicated by the progressive decrease in the size of the AII-induced Ca2+ transient, showed a rapid initial (t1/2 = 1.7 min) component that accounted for about 80% of the response and a slowly decreasing phase with t1/2 = 112 min. The latter thapsigargin-resistant component was abolished by the removal of extracellular Ca2+. Pretreatment with AII dose-dependently attenuated but did not abolish the subsequent Ca2+ response to thapsigargin and also increased the rate of the Ca2+ rise induced by thapsigargin. In bovine adrenocortical microsomes, thapsigargin inhibited the ATP-dependent filling of Ca2+ pools and caused a dose-dependent rise in extravesicular Ca2+ levels when added to previously loaded microsomes. The thapsigargin-releasable Ca2+ pool in adrenal microsomes was larger than the inositol 1,4,5-trisphosphate (Ins(1,4,5)P3)-sensitive Ca2+ pool but only slightly greater than the GTP-releasable pool. Ins(1,4,5)P3-induced Ca2+ release was reduced markedly when ATP-dependent Ca2+ loading of the microsomes was prevented by prior addition of thapsigargin. However, the subsequent Ca2+ response to Ins(1,4,5)P3 was consistently better preserved after the addition of thapsigargin to microsomes preloaded with Ca2+. This difference suggests that although Ca2+ uptake by the Ins(1,4,5)P3-responsive pool is also sensitive to thapsigargin, once filled, this pool shows a slower passive leakage than other thapsigargin-sensitive pools. These findings indicate that thapsigargin increases [Ca2+]i by inhibiting Ca2+ uptake into multiple intracellular Ca2+ pools and by also promoting entry of extracellular Ca2+.(ABSTRACT TRUNCATED AT 400 WORDS)

8-Diethylamino-octyl-3,4,5-trimethoxybenzoate, a calcium store blocker, increases calcium influx, inhibits alpha-1 adrenergic receptor calcium mobilization, and alters iodide transport in FRTL-5 rat thyroid cells.

8-Diethylamino-octyl-3,4,5-trimethoxybenzoate (TMB-8) is known to inhibit mobilization of calcium from intracellular stores but, more recently, other cellular effects have been described. In the present study, the effects of TMB-8 on cytosolic free calcium [Ca2+]i levels in FRTL-5 rat thyroid cells were determined using the fluorescent dye, Indo-1. TMB-8 increased [Ca2+]i in a dose-dependent manner, with a maximum rise from 120 +/- 7 nM to 229 +/- 16 nM (90 +/- 5% increase) at 5 x 10(-4) M. This effect was considerably reduced in Ca2+ free buffer, demonstrating a dependency upon extracellular calcium influx but not upon membrane potential and which did not involve the Na+/Ca2+ exchanger. In Ca2+ free buffer TMB-8, at a dose which did not affect [Ca2+]i, completely prevented norepinephrine (10(-5) M) from mobilizing intracellular Ca2+. To determine whether TMB-8 affected differentiated functions, iodide uptake and efflux studies were performed with 125I. TMB-8 (10(-4) M) inhibited iodide uptake by approximately 40% without affecting efflux. At 10(-3) M TMB-8, efflux was also enhanced. These studies demonstrate that TMB-8 has at least two effects on [Ca2+]i, promoting calcium influx and preventing alpha-1 adrenergic mobilization from intracellular stores. TMB-8 also has multiple effects on 125I transport, both inhibiting influx and increasing efflux. The results emphasize the importance of characterizing the behavior of this compound in any cell system before using it as a biological probe.

Interaction between serotonin and other regulators of aldosterone secretion in rat adrenal glomerulosa cells.

Although serotonin (5HT) is a recognized stimulator of aldosterone secretion in vivo and in vitro, its physiological role as a regulator of mineralocorticoid secretion and its mechanism of action in the adrenal glomerulosa have not been elucidated. To address these questions we studied the interaction of 5HT with other aldosterone regulators in isolated rat adrenal glomerulosa cells. 5HT stimulated aldosterone production 14-fold, with an ED50 of 20 +/- 5 nM, and stimulation was maximal at 0.8 microM. The stimulation of aldosterone production by 5HT was accompanied by a 5-fold increase in cAMP production, with an ED50 of 1 microM. Threshold levels of 5HT (1 nM) potentiated the effect of submaximal concentrations of angiotensin-II (AII), decreasing the ED50 from 1.3 to 0.46 nM and increasing the maximum response in an additive manner. In contrast, the stimulatory effect of 5HT was purely additive to that of submaximal ACTH concentrations. 5HT had no effect on aldosterone secretion stimulated by maximal ACTH concentrations, despite full additivity on cAMP accumulation. Stimulations of steroidogenesis by potassium and 5HT were fully additive at submaximal concentrations, but only partially additive at-maximal levels. To determine the mechanism of the synergistic effects of AII and 5HT, we analyzed the interaction of both stimuli on cAMP accumulation, intracellular calcium, and inositol phosphate formation. Consistent with the inhibitory effect of AII on adenylate cyclase, in the presence of AII the stimulation of cAMP by 5HT was reduced by 18 +/- 3%. 5HT alone had no effect on cytosolic calcium, but significantly enhanced the peak and later phases of the AII-stimulated increase (P less than 0.005). This effect of 5HT was due to calcium influx and not to release from intracellular pools, as shown by suppression of the potentiation in the absence of extracellular calcium and the lack of effect of 5HT on basal or AII-stimulated inositol phosphate formation. The ability of low concentrations of 5HT to potentiate the stimulatory effect of AII on aldosterone secretion suggests that under some physiological conditions, 5HT may play a role in regulating the adrenal sensitivity to AII.

Na(+)-ionophore, monensin-induced rise in cytoplasmic free calcium depends on the presence of extracellular calcium in FRTL-5 rat thyroid cells.

Calcium is an important regulator of cell function, and may be influenced by the intracellular sodium content. In the present study, the Na(+)-ionophore, monensin, was used to investigate the interrelationship between changes in intracellular Na+ concentration ([Na+]i) and elevation of cytosolic Ca2+ concentration ([Ca2+]i) in FRTL-5 thyroid cells. Cytoplasmic Ca2+ levels were measured using the fluorescent dye, indo-1. Monensin induced a dose-dependent increase in [Ca2+]i in FRTL-5 cells. Inhibitors of intracellular Ca2+ release, TMB-8 and ryanodine, were unable to prevent the monensin effect on [Ca2+]i. The alpha 1-receptor antagonist, prazosin, did not block the monensin-stimulated increase in [Ca2+]i. In the absence of extracellular calcium there was a marked diminution in the monensin effect on [Ca2+]i, yet calcium channel antagonists (nifedipine, diltiazem and verapamil) did not inhibit the response. Replacement of Na+ by choline chloride in the medium depressed the monensin-evoked rise in [Ca2+]i by up to 84%. Furthermore, addition of the Na(+)-channel agonist, veratridine, elicited an increase in [Ca2+]i, even though less dramatic than that caused by monensin. Ouabain increased the resting cytosolic Ca2+ concentration as well as the magnitude of the monensin effect on [Ca2+]i. The absence of any effect on the Na(+)-ionophore evoked increase in [Ca2+]i upon addition of tetrodotoxin (TTX) excluded a possible involvement of TTX-sensitive Na+ channels. These data show that the rise in [Ca2+]i induced by increasing [Na+]i is largely dependent on both external Na+ and Ca2+. Calcium entry appears not to involve voltage-dependent or alpha 1-receptor sensitive Ca2+ channels, but may result from activation of an Na(+)-Ca2+ exchange system.