Phosphorylation processes mediate rapid changes of brain aromatase activity

Abstract

The enzyme aromatase (also called estrogen synthase) that catalyzes the transformation of testosterone (T) into estradiol plays a key limiting role in the action of T on many aspects of reproduction. The distribution and regulation of aromatase in the quail brain has been studied by radioenzyme assays on microdissected brain areas, immunocytochemistry, RT-PCR and in situ hybridization. High levels of aromatase activity (AA) characterize the sexually dimorphic, steroid-sensitive medial preoptic nucleus (POM), a critical site of T action and aromatization for the activation of male sexual behavior. The boundaries of the POM are clearly outlined by a dense population of aromatase-containing cells as visualized by both immunocytochemistry and in situ hybridization histochemistry. Aromatase synthesis in the POM is controlled by T and its metabolite estradiol, but estradiol receptors alpha (ERα) are not normally co-localized with aromatase in this brain area. Estradiol receptor beta (ERβ) has been recently cloned in quail and localized in POM but we do not yet know whether ERβ occurs in aromatase cells. It is therefore not known whether estrogens regulate aromatase synthesis directly or by affecting different inputs to aromatase cells as is the case with the gonadotropin releasing hormone neurons. The presence of aromatase in presynaptic boutons suggests that locally formed estrogens may exert part of their effects by non-genomic mechanisms at the membrane level. Rapid effects of estrogens in the brain that presumably take place at the neuronal membrane level have been described in other species. If fast transduction mechanisms for estrogen are available at the membrane level, this will not necessarily result in rapid changes in brain function if the availability of the ligand does not also change rapidly. We demonstrate here that AA in hypothalamic homogenates is rapidly down-regulated by exposure to conditions that enhance protein phosphorylation (addition of Ca 2+, Mg 2+, ATP). This inhibition is blocked by kinase inhibitors which supports the notion that phosphorylation processes are involved. A rapid (within minutes) and reversible regulation of AA is also observed in hypothalamic explants incubated in vitro and exposed to high Ca 2+ levels (K +-induced depolarization, treatment by thapsigargin, by kainate, AMPA or NMDA). The local production and availability of estrogens in the brain can therefore be rapidly changed by Ca 2+ based on variation in neurotransmitter activity. Locally-produced estrogens are as a consequence available for non-genomic regulation of neuronal physiology in a manner more akin to the action of a neuropeptide/neurotransmitter than previously thought.