Transforming growth factor- alpha 's effects on astroglial-cholinergic cell interactions in the medial septal area in vitro are mediated by alpha sub(2)-macroglobulin

We reported previously that two epidermal growth factor receptor ligands, epidermal growth factor and transforming growth factor- alpha , inhibit medial septal cholinergic cell phenotypic expression (choline acetyltransferase and acetylcholinesterase activities) in vitro indirectly via (a) soluble molecule(s) released from astrocytes. In the present study, we found that this response to transforming growth factor- alpha is mediated, for the most part, by alpha sub(2)-macroglobulin, a potent protease inhibitor with a wide spectrum of biological activities. In this regard, the effects of transforming growth factor- alpha on cholinergic cells can be blocked with immunoneutralizing antibodies raised against alpha sub(2)-macroglobulin. Furthermore, western blot analysis reveals that although alpha sub(2)-macroglobulin is present in conditioned media from control septal cultures, it is more abundant in those treated with transforming growth factor- alpha . In addition, exogenous alpha sub(2)-macroglobulin, both in its native and trypsin-activated forms, can mimic transforming growth factor- alpha 's effects on septal cholinergic cell expression. However, while the native antiprotease can slightly but significantly decrease choline acetyltransferase activity, trypsin-activated alpha sub(2)-macroglobulin, in the nanomolar range, induces as marked a decrease in this enzyme activity as that noted with transforming growth factor- alpha . Furthermore, trypsin-activated alpha sub(2)-macroglobulin, like epidermal growth factor /transforming growth factor- alpha , decreases choline acetyltransferase activity by arresting its spontaneous increase that occurs with time in culture, does so in a reversible manner and is not neurotoxic. In addition, trypsin-activated alpha sub(2)-macroglobulin, in the nanomolar range, can affect choline acetyltransferase in a dual manner, up-regulating it at low concentrations while down-regulating it at higher ones. These responses are identical in mixed neuronal-glial and pure neuronal septal cultures. Furthermore, when concentrations of trypsin-activated alpha sub(2)-macroglobulin, which alone decrease choline acetyltransferase, are added simultaneously with nerve growth factor, they serve to potentiate the nerve growth factor-induced increase in enzymatic activity. As GABAergic cell expression is not affected by alpha sub(2)-macroglobulin, it appears that the effects of this protease inhibitor on medial septal neuronal expression are neur