Folliculostellate (FS) cells within the anterior pituitary gland produce cytokines and growth factors which are regulated by molecules produced by neighbouring endocrine cells. There are many factors that could potentially be responsible for this intercellular communication that is mediated via connexins (Cx) and gap-junctions. We thus investigated the effect of co-culturing lactotrophs (MMQ) and FS cells (TtT/GF) on Cx43 expression. Our in vitro model system consists of culturing TtT/GF cells in replicate multiwells and MMQ cells in well inserts. This system allows cellular products that are secreted (over 24 hrs) from MMQ cells to affect the activity of TtT/GF cells.
Western blotting analysis of Cx43 expression in TtT/GF cells showed that both the phosphorylated and non-phosphorylated forms increased in a manner that was directly related to the number of MMQ cells plated out. Parallel findings were obtained when adenosine or the universal adenosine receptor agonist (NECA) were added directly to MMQ cells. NECA stimulated Cx43 expression with an EC50=0.1 μM and reached a plateau within 2 hr. Analysis of cell-conditioned media by reverse-phase fluorescent HPLC analysis shows that MMQ (and GH3) cells spontaneously secrete adenosine (34 μM/hr) and are able to dephosphorylate exogenously added fluorescent AMP to adenosine with a half life of 1 hr.
In summary our findings show that MMQ cells spontaneously secrete adenosine to levels that can modulate Cx43 expression and potentially influence gap-junction communication. These data suggest that the functional activity of FS cells may be dependent on the metabolic activity of adjoining endocrine cells.