Carbachol increases intracellular free calcium concentrations in human granulosa-lutein cells

Mayerhofer, Artur; Föhr, Karl J.; Sterzik, K.; Gratzl, Manfred

doi:10.1677/joe.0.1350153

Cited by 37 publications

(42 citation statements)

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“…Luck (1990) reported the stimulation in the production of steroids in bovine granulosa cells by cholinergic effect in ovary whereas Morley et al (1992) did not find any increase in the progesterone production in chicken granulosa cells. Mayerhofer et al (1992) did not find modifications in the liberation of progesterone in human granulosa cells culture by acetylcholine effect. On the contrary, Bodis et al (1993) have stated that the cholinergic neurotransmission may have a physiological significance in the intraovarian regulatory pathways.…”

Section: Discussionmentioning

confidence: 70%

The cholinergic influence on the mesenteric ganglion affects the liberation of ovarian steroids and nitric oxide in oestrus day rats: characterization of an ex vivo system

Orozco¹,

Sosa²,

Fillipa³

et al. 2006

Journal of Endocrinology

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The axons that constitute the ovarian nervous plexus originate mostly in the principal neurons of the superior mesenteric ganglion (SMG) that is part of the sympathetic ganglionic chain and exhibits cholinergic receptors. In order to observe the effect of acetylcholine, the main neurotransmitter in the ganglionic transmission, the purpose of the present work was: first, to standardize an integrated ex vivo superior mesenteric ganglion-ovarian nervous plexus-ovary (SMG-ONP-O) system in oestrus day rats; secondly, to determine if the ganglionic cholinergic stimulus modifies the release of nitric oxide and steroids in the ovary compartment in the absence of humoral factors; and thirdly, to investigate if there are differences in the responses between the left and right ovaries caused by the neural stimulus. The ex vivo experimental left and right systems were developed and standardized. The systems were incubated in Krebs-Ringer phosphate buffer in a Dubnoff metabolic shaker. The progesterone release was determined to standardize the incubation times, obtaining different responses between the left and right systems, which shows that both systems have their own autonomic tone. Non-specific stimulation with KCl in the ganglion compartment provoked different responses in terms of release of progesterone and oestradiol. Progesterone decreased in the left and right systems.However, oestradiol diminished at short times and increased at 60 and 120 min in the left ovary, whereas it increases at 30 and 60 min in the right ovary. These different responses show the sensitivity and viability of both systems. When acetylcholine was used in the ganglion compartment, the release of nitric oxide, progesterone, androstenedione and oestradiol was evaluated. The liberation of nitrite increased at 15, 30 and 60 min in the left system and decreased in the right system at 120 min. Progesterone showed a decrease in its release at 15, 30 and 120 min and androstenedione at 15 min in the left ovary compartment. In the right ovary, only progesterone decreased in relation to the control at 120 min while androstenedione did not show significant changes. Oestradiol showed an increase in the left ovary compartment at all the studied times, while in the right ovary it did not show any changes. These results indicate that the neural stimulus from the superior mesenteric ganglion through the ovarian nervous plexus is one of the factors modulating the secretory activity of the ovarian steroids and nitric oxide. The system is viable and also shows a different sensitivity of the left ovary in relation to the right one at least in this cycle stage, characterized by marked irrigation and profound structural changes in the ovary.

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Section: Discussionmentioning

confidence: 70%

The cholinergic influence on the mesenteric ganglion affects the liberation of ovarian steroids and nitric oxide in oestrus day rats: characterization of an ex vivo system

Orozco¹,

Sosa²,

Fillipa³

et al. 2006

Journal of Endocrinology

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“…Even in the absence of extracellular Ca2+, oxytocin caused a Ca2+ transient in responsive cells, indicating the involvement of intracellular Ca2+ stores. Additional evidence for the nature of the intracellular source of Ca2+ is provided by experiments with ionomycin (15,29,30). Thus, depletion of intracellular Ca2+ stores (by the ionophore ionomycin) completely abolished the response of the cells upon oxytocin stimulation.…”

Section: Discussionmentioning

confidence: 99%

“…The procedure has been described previously in detail (15,16 Germany), as described previously (15). Ratios of the recordings at 340 and 380 nm were determined as previously described (20).…”

Section: Methodsmentioning

confidence: 99%

“…The total volume per dish was 3 mL. After 6 h, the supernatants were collected and frozen until measurement of progesterone using routine RIA protocols at the Universitatsfrauenklinik Ulm, as previously described (15,16 …”

Section: Measurementsmentioning

confidence: 99%

“…Solution A was modified by adding 1 mmol/L EGTA instead of CaCh to perform measurements in the absence of extracellular Ca2+. Fluorescence measurements were made with the Zeiss Microscope Photometer System (FFP, Zeiss, Oberkochen, Germany), as described previously (15). Ratios of the recordings at 340 and 380 nm were determined as previously described (20).…”

Section: Calcium Measurementsmentioning

confidence: 99%

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Effect of oxytocin on free intracellular Ca2+ levels and progesterone release by human granulosa-lutein cells.

Mayerhofer

Sterzik

Link

et al. 1993

The Journal of Clinical Endocrinology & Metabolism

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Oxytocin and its receptor are found in the corpus luteum in a variety of species, including the human.In the present study we used fura- T HE SYNTHESIS of oxytocin in the corpus luteum of the ovary is well established and occurs in a variety of species (l-3; for review, see Ref. 4), including the human (5-8). Moreover, indirect evidence for the presence of oxytocin receptors in human granulosa-lutein cells was provided by a study that showed inhibitory effects of oxytocin on FSHstimulated estradiol secretion (9). Using microdialyzed human corpora lutea, paracrine effects of oxytocin on steroid production were reported. Oxytocin stimulated both progesterone and estradiol release in the young human corpora lutea examined (10). Besides these reports, which give indirect evidence for functional oxytocin receptors in the human ovary, more recently their presence was proven by the detection of oxytocin receptor mRNA in the human ovary (11). However, the signal transduction pathway of the oxytocin receptor in the human corpus luteum has not been examined, and there is no information available concerning whether the expression of oxytocin receptors changes during differentiation. We, therefore, tried to address these points in the present study. We investigated whether activation of oxyto- tin receptors can cause elevations of intracellular free calcium concentrations in human granulosa-lutein cells in a manner similar to that we reported previously in cultured human myometrial cells (12) and rat pituitary corticotrophs (13,14). In addition, we examined the role of oxytocin in progesterone release, and the effect of hCG exerted on the release of oxytocin by cultured human granulosa-lutein cells. Materials and Methods Materials ABSTRACTOxytocin and its receptor are found in the corpus luteum in a variety of species, including the human. In the present study we used fura-2 microfluorimetry to investigate whether activation of the oxytocin receptor of cultured human granulosa-lutein cells causes intracellular calcium (Ca 2 +) signals and affects progesterone release. Although after 1 day in culture, ceHs were not responsive to oxytocin, the number of responsive cells increased steadily during the first 3 days in culture, reaching a maximum on days 4 and 5 (59-66%) and then declined again until day 8. Effective oxytocin concentrations were apparently independent of the culture day, and concentrations as low as 10 nmoljL increased intracellular free Ca 2 + levels from 70-140 nmol/L (basal levels) to maximal peak levels of 800 nmoljL. The oxytocin-induced Ca 2 + signal was not affected by removal of extraceHular Ca 2 + with EGTA. Moreover, depletion of intracellular Ca 2 + stores by ionomycin treatment rendered the cells unresponsive to oxytocin, pointing also at the intraceHular source of the oxytocin-inducible Ca

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Ovarian acetylcholine and muscarinic receptors: Hints of a novel intrinsic ovarian regulatory system

Mayerhofer

Fritz

2002

Microscopy Res & Technique

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More than two decades ago, the degrading enzyme of the neurotransmitter acetylcholine (ACH) was reported in nerve fibers of the rat ovary. Subsequently, it was assumed that ACH is a neurotransmitter of ovarian nerves, although the sole presence of the degrading enzyme, ACH-esterase, does not allow such a conclusion. That ACH may be involved in the complex regulation of ovarian functions, including hormone production, was indicated by studies using, for example, granulosa cells (GCs). The lack of detailed information about both source(s) and functions of ACH in the ovary prompted us to examine sites of ovarian ACH-synthesis and ACH-receptor-bearing target cells. We also started to identify functions of ACH in cultured human GCs. While ovarian innervation and recently described neuron-like cells of the ovary were not immunoreactive for the ACH-synthesizing enzyme, choline-acetyl transferase (CHAT), we found immunoreactivity in GCs of rodents and primates. Isolated human and rat GCs produced ACH and contained the vesicular ACH transporter (VACHT). These results indicate that endocrine GCs are an unexpected non-neuronal source of ACH in the ovary. Moreover, these cells and GCs in vivo contain ACH-receptors of the muscarinic subtype (MR), namely M1R and M5R. In contrast, oocytes express M3R. MR of human GCs are functional and cholinergic stimulation is linked to rapid increases in intracellular Ca(++) levels. M1/5R activation also led to increased cell proliferation of human GCs in vitro and this stimulatory effect was found to be associated with rapid disruption of gap junction communication. Ongoing studies begin to identify regulation of ion channels and altered gene expression as consequences of MR stimulation. Thus, our results outline first details of an unexpected intraovarian, non-neuronal cholinergic system, and suggest that it may be involved in the regulation of cell proliferation in the ovary.

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Carbachol increases intracellular free calcium concentrations in human granulosa-lutein cells

Cited by 37 publications

References 8 publications

The cholinergic influence on the mesenteric ganglion affects the liberation of ovarian steroids and nitric oxide in oestrus day rats: characterization of an ex vivo system

The cholinergic influence on the mesenteric ganglion affects the liberation of ovarian steroids and nitric oxide in oestrus day rats: characterization of an ex vivo system

Effect of oxytocin on free intracellular Ca2+ levels and progesterone release by human granulosa-lutein cells.

Ovarian acetylcholine and muscarinic receptors: Hints of a novel intrinsic ovarian regulatory system

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