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Remote Neuronal Activation Coupled with Automated Blood Sampling to Induce and Measure Circulating Luteinizing Hormone in Mice
Published on: August 25, 2023
Luteinizing and human chorionic gonadotropin hormones increase intercellular communication and gap junctions in
E M Pérez-Armendariz1, J Luna, C Miranda
1Department of Neuroscience, Albert Einstein College of Medicine, Bronx, NY.
Insights
Luteinizing hormone (LH) and human chorionic gonadotropin (hCG) maintain intercellular communication and gap junctions in mouse Leydig cells. These hormones, via cAMP, prevent the age-related decline in cell communication and connexin 43 expression.
Area of Science:
- Reproductive biology
- Cell biology
- Endocrinology
Background:
- Leydig cells are crucial for testosterone production in the testes.
- Intercellular communication (ic) and gap junctions (GJs) play vital roles in testicular function.
- The influence of LH and hCG on GJ-mediated ic in Leydig cells requires further elucidation.
Purpose of the Study:
- To investigate the effects of LH and hCG on gap junctions and intercellular communication in mouse Leydig cells.
- To determine if cAMP mediates the effects of LH and hCG on Leydig cell function.
- To assess the impact of these hormones on testosterone secretion.
Main Methods:
- Primary Leydig cell cultures were treated with LH, hCG, or dibutiryl-cAMP (db-cAMP).
- Intercellular communication was assessed using Lucifer yellow microinjection.
- Gap junction expression was evaluated via immunofluorescent labeling of connexin 43 (Cx43).
- Testosterone levels were quantified using radioimmunoassay.
Main Results:
- LH and hCG treatments increased testosterone secretion at all time points.
- In aged cultures (24-36h), intercellular communication and Cx43 expression decreased significantly.
- LH, hCG, and db-cAMP prevented the decline in intercellular communication and Cx43 expression in aged cultures.
- Immunoblotting confirmed increased Cx43 protein levels in hormone-treated cells.
Conclusions:
- LH and hCG modulate connexin 43 expression and intercellular communication in mouse Leydig cells.
- The effects of LH and hCG appear to be mediated by the cAMP signaling pathway.
- Maintaining intercellular communication is crucial for Leydig cell function and steroidogenesis.
Abstract:
The effect of luteinizing (LH) and human chorionic gonadotropin (hCG) hormones on gap junctions (Gjs) and intercellular communication (ic) was evaluated in Leydig (interstitial) cells from mouse testes. Cell cultures enriched in Leydig cells were studied under control conditions and when maintained in the presence of 100 ng/mL LH, 10 ng/mL hCG, or 1 mM dibutiryl-cAMP (db-cAMP), for 8, 24, and 36 h. To monitor the extent of ic, Lucifer yellow (LY) was injected through a patch pipet into one cell of-small cell aggregates (6-10), and its transfer was evaluated using fluorescent microscopy. The expression of GJs was monitored using immunofluorescent (IF) labeling of connexin 43 (Cx43) with a specific antibody. Testosterone secretion was determined by radioimmunoassay. At all culture times, testosterone levels in the medium were higher in treated than in control cell cultures. In cell cultures of 8 h, LY transferred to most of the neighboring cells (93%) and cell membrane appositions showed abundant Cx43; no difference was found between control and treated cells. In contrast, in control cell cultures of 24 and 36 h, LY transferred to a reduced fraction of neighboring cells (46 and 21%, respectively) and Cx43 labeling was markedly decreased. Addition of LH, hCG, or db-cAMP, to cell cultures for 24 and 36 h completely prevented the decrease in ic and Cx43 expression. Immunoblot studies, from total protein homogenates of cell cultures of 36 h, showed that relative levels of 40- and 43-kDa bands, characteristic of Cx43, were higher in treated than in control cells. These results demonstrate that the expression of Cx43 and ic in Leydig cells is modulated by LH and hCG, and suggest that their effect is mediated by the second messenger of these hormones, cAMP.
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