Acute Effects of Adenosine Triphosphates, Cyclic 3′,5′-Adenosine Monophosphates, and Follicle-Stimulating Hormone on Cytosolic Calcium Level in Cultured Immature Rat Sertoli Cells

Abstract

The ability of ATP and FSH to induce intracellular calcium [Ca(2+)](i) changes in Sertoli cells is imperfectly understood and reports are conflicting. We have applied the single-cell microfluorometry technique with the calcium probe indo-1 to investigate [Ca(2+)](i) in individual cultured Sertoli cells. When cells were exposed to ATP, cAMP, and FSH, a fast and biphasic increase in [Ca(2+)](i) was obtained in 100%, 70%, and 56% of cells, respectively. Caffeine did not activate Ca(2+) mobilization, while thapsigargin suppressed the peak response. External calcium free-EGTA buffer suppressed the plateau phase, while blockers of voltage-operated Ca(2+) channels did not abolish the response to cAMP and ATP. We conclude that the three messengers mobilized Ca(2+) from intracellular thapsigargin-sensitive stores, which induced a subsequent Ca(2+) influx from the extracellular medium by a voltage-independent Ca(2+) entry. The well-documented mechanisms by which these messengers act on cells support the idea that they release Ca(2+) from smooth endoplasmic reticulum by two different pathways, or that FSH and cAMP first release ATP, which then acts on cells. Among the cells, 77% and 80% responded, respectively, to FSH and cAMP by a delayed long-lasting decrease in [Ca(2+)](i) that was never recorded in the presence of ATP. This suggests that FSH and cAMP also promote a slow redistribution of [Ca(2+)](i) from the exchangeable pool to the bound nonexchangeable pools. Involvement of voltage-operated and voltage-independent calcium channels in the response of Sertoli cells to ATP, FSH, and cAMP is discussed.

Mechanisms involved in the homologous down-regulation of transcription of the follicle-stimulating hormone receptor gene in Sertoli cells

“…The biological events controlled by this endocrine mechanism result from the interaction of follicle-stimulating hormone (FSH) with the FSH receptor and the subsequent transduction of molecular information across the membrane leading to the production of second messengers such as cAMP (Means et al, 1980) and Ca 2 + (Chaudhary et al, 1996;Lalevee et al, 1999) and changes in gene expression.…”

Expression of Nucleotide P2X Receptor Subtypes during Spermatogenesis in the Adult Rat Testis

“…A cell-type-and maturation stage-specific receptor expression, as shown for P2X receptors in this study, could account for coordinated purinergic signalling within one homogeneous group, separating it (in respect of purinergic signalling) from a different group not expressing P2X receptors. Lalevée et al [1999] speculated upon a different pathway for ATP release, experimenting with immature rat Sertoli cell cultures. These authors describe a possible ATP release from Sertoli cells after stimulation with follicle-stimulating hormone (FSH), and release of ATP by germ cells.…”

“…Profound changes take place at stage VIII, as the mature generation of sperm is released [Leblond and Clermont, 1952] and Sertoli cells change their hormone responsiveness at this stage, especially their responsiveness to FSH [Parvinen et al, 1980]. One might speculate that this change in FSH responsiveness of Sertoli cells could account for changes in the magnitude of ATP secretion from Sertoli cells, according to the mechanism suggested by Lalevée et al [1999]. In the subsequent stages (IX-XIV) P2X 2 and P2X 3 receptors were absent from the tubules but P2X 5 receptors were detected through stages X-XIII.…”

Calcium confusion--is the variability in calcium response by Sertoli cells to specific hormones meaningful or simply redundant?

“…Differences in labelling or sub-cellular distribution of the calcium-sensitive fluorescent dye, backtransport or leakage of the dye after stimulation (especially in freshly isolated cells) and photobleaching could mask the authentic calcium response. However, in several instances cells did not show a calcium response to the specific hormone applied whereas, in the same cells, calcium responses could be shown after addition of potassium, foetal calf serum or ATP (D'Agostino et al 1993, Lalevee et al 1999. Thus technical problems seem to be of less importance and the variations in hormone-dependent calcium responses appear to depend more on the properties of particular receptors or cellular set points.…”

“…The suggestion has been made that the basal calcium concentration could be an important factor for the direction of the response after hormonal stimulation (Lalevee et al 1999), but it is hard to understand why basal calcium concentrations vary between similar experiments within a single laboratory. We have made many attempts to understand the origin of these calcium variations by making numerous alterations to the experimental conditions.…”