Progesterone Antagonism of Neurite Outgrowth Depends on Microglial Activation via Pgrmc1/S2R

Abstract

Neuronal plasticity is regulated by the ovarian steroids estradiol (E2) and progesterone (P4) in many normal brain functions, as well as in acute response to injury and chronic neurodegenerative disease. In a female rat model of axotomy, the E2-dependent compensatory neuronal sprouting is antagonized by P4. To resolve complex glial-neuronal cell interactions, we used the "woundingin-a-dish" model of neurons cocultured with astrocytes or mixed glia (microglia to astrocytes, 1:3). Although both astrocytes and mixed glia supported E2-enhanced neurite outgrowth, P4 antagonized E2-induced neurite outgrowth only with mixed glia, but not astrocytes alone. We now show that P4-E2 antagonism of neurite outgrowth is mediated by microglial expression of progesterone receptor (Pgr) membrane component 1 (Pgrmc1)/S2R, a putative nonclassical Pgr mediator with multiple functions. The P4-E2 antagonism of neurite outgrowth was restored by add-back of microglia to astrocyte-neuron cocultures. Because microglia do not express the classical Pgr, we examined the role of Pgrmc1, which is expressed in microglia in vitro and in vivo. Knockdown by siRNA-Pgrmc1 in microglia before add-back to astrocyte-neuron cocultures suppressed the P4-E2 antagonism of neurite outgrowth. Conditioned media from microglia restored the P4-E2 activity, but only if microglia were activated by lipopolysaccharide or by wounding. Moreover, the microglial activation was blocked by Pgmrc1-siRNA knockdown. These findings explain why nonwounded cultures without microglial activation lack P4 antagonism of E2-induced neurite outgrowth. We suggest that microglial activation may influence brain responses to exogenous P4, which is a prospective therapy in traumatic brain injury. (Endocrinology 154: 2468 -2480, 2013) C ross talk between estradiol (E2) and progesterone (P4) operates throughout the female reproductive system in cycles of E2-dependent cell growth and P4-dependent cell regression. However, little is known of the brain cell interactions that enable "P4-E2 antagonism" in synaptic plasticity. In neuronal responses to injury that involve P4-E2 antagonism, we describe a novel role of microglial activation that is mediated by progesterone receptor (Pgr) membrane component 1 (Pgrmc1)/S2R, a nonclassical Pgr of emerging significance to brain neuronal functions and innate immunity.In the hippocampus, synaptic remodeling during the rat estrous cycle is driven by serial elevations of E2 and P4 (1). Neuronal sprouting in response to axotomy is also modulated by P4-E2 antagonism. In an adult rat model of entorhinal cortex lesions that axotomize the perforant pathway to the hippocampus, the ensuing compensatory neurite outgrowth into the dentate gyrus molecular layer was stimulated by E2 and blocked by concurrent administration of E2ϩP4 (2). Astrocyte and microglial activation in the deafferented zone was correspondingly modulated by E2 and P4. An unexpected role of microglia in P4-E2 antagonism of neurite outgrowth was identified with the "wounding-in-a-dish" model of ax...

Pgrmc1: new roles in the microglial mediation of progesterone-antagonism of estradiol-dependent neurite sprouting and in microglial activation

“…The two classic nuclear gene transcription factors PR-A and PR-B are alternate transcripts of Pgr , are widely expressed in neurons throughout the adult rat brain including cerebral cortex, hippocampus, and hypothalamus (Intlekofer and Petersen, 2011; Bali et al, 2012). We detected PRs in astrocytes, but not in microglia of adult rat hippocampus ( in situ hybridization and immunocytochemistry), and in primary glial cultures from neonatal cortex by rtPCR and Western blots (Figure 1) (Bali et al, 2013). These findings confirm observations of the absence of Pgr in ex vivo fluorescence-sorted (FACS) adult mouse brain microglia (Sierra et al, 2008).…”

“…Unlike Pgr, Pgrmc1 is uniformly expressed by all hippocampal neurons (Bali et al, 2012). Although initially characterized as a membrane receptor, Pgrmc1 can reside in the cell nucleus (Peluso et al, 2010; Bali et al, 2013). We also detected other membrane-associated PRs, mPRα, mPRβ, and mPRγ, in astrocytes and microglia (rtPCR, DNA sequence confirmed; unpublished data).…”

“…A caveat is that the 4 h mechanical shaking process to remove microglia could alter glial responses, e.g., hydrodynamic forces induce astrocyte aromatase (Gatson et al, 2011). The role of microglia was shown in a new protocol to add-back microglia, which restored the P4-E2 antagonism of neurite outgrowth (Bali et al, 2013). …”

“…Glial activation by injury or LPS and the SA effects were blocked by Pgrmc1 knockdown. Modified from Bali et al (2013). …”

“…Other family members include Pgrmc2, neudesin, and neuferricin, which are distinct from the classical progesterone receptor (PR) transcription factors and from the large family of progestin/adipoQ membrane receptors that include mPRα, mPRβ, and mPRγ. We recently discovered two new roles of Pgrmc1 in microglial activation and in the microglial mediation of ovarian steroid effects on neurite sprouting (Bali et al, 2013). …”