Women are more susceptible than men to develop anxiety disorders, however, the mechanisms involved are still unclear. In this study, we investigated the role of group I metabotropic glutamate receptors (mGluRs), a target for anxiety disorders, and whether estradiol may modulate conflict-based anxiety in female rats by using the Vogel Conflict Test (VCT). We used ovariectomized female rats with high (OVX + EB) and low (OVX) estradiol levels and intact male rats to evaluate sex differences. Infusion of (S)-3,5-Dihydroxyphenylglycine (DHPG), a group I mGluR agonist, into the basolateral amygdala, a region involved in anxiety-responses, statistically increased the number of shocks in OVX, but not OVX + EB female rats at 0.1, nor at 1.0 μM. In contrast, DHPG statistically decreased the number of shocks in male rats at 1.0 μM only. DHPG (0.1 μM) increased the number of recoveries in OVX, but not OVX + EB or male rats. Sex differences were detected for the number of shocks, recoveries and punished licks, where female rats displayed more conflict than male rats. Western blot analyses showed that protein expression of mGluR1, but not mGluR5 was higher in OVX + EB > OVX > male rats in the amygdala, whereas no significant differences were detected in the hippocampus, olfactory bulb and/or the periaqueductal gray. Therefore, DHPG produced paradoxical effects that are sex dependent; producing anxiolytic-like effects in female rats, while anxiogenic-like effects in male rats according to the VCT. These results highlight the importance of including female experimental models to underpin the neural circuitry of anxiety according to sex and for the screening of novel anxiolytic compounds.

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Sex differences in the manifestation, prevalence, and progression of dopamine‐related diseases have been documented, suggesting a role of gonadal hormones (Bodzean et al., 2014; Kokras and Dalla, 2014). Although dopamine neurons are sensitive to estradiol (E2), main estrogen metabolite in women, it is uncertain how estradiol affects the function and survival of dopamine neurons. Eight metabotropic glutamate receptor subtypes (mGluR1‐8) have been identified and classified in three different groups according to the receptor signal transduction, pharmacology and amino acid sequence homology. mGluR subtypes are important modulators of cell proliferation (Plenz and Kitaj 1998), neuronal excitability and neurotransmitter release (Meltzer et al. 1997; Mercuri et al. 1993). The objective of our research was to study the effects of E2 on the expression of group I (mGluR1 and 5) and group II (mGluR2 and 3) mGluRs. We hypothesized that E2 increases the protein expression of group I mGluRs without affecting the expression of group II mGluRs. To test our hypothesis cultures of embryonic mesencephalic dopamine neurons were treated with either cyclodextrin or E2 at different concentrations (10nM and 10μM). Immunocytochemistry experiments showed the expression of estrogen receptors alpha (ERα), mGluR1, and mGluR2/3 subtypes in our experimental cellular model. Western bot analyses demonstrated that E2 treatment upregulates the expression of mGluR1, a group I mGluR subtype, but not group II mGluRs subtypes. Estradiol's effect was concentration dependent producing mGluR1 upregulation at 10μM but not at 10nM (F(2,9)=4.03, p=0.05). Changes in mGluRs protein expression by differences in estradiol levels may underlay variations in cells excitability and function, inducing sex differences in the neurophysiology of dopamine neurons.Support or Funding InformationThis research was supported by the Deanship of Academic Affairs of the University of Puerto Rico, Cayey Campus (FIDI‐UPR CAYEY 2017–2018) and in part by Puerto Rico Center for Environmental Neuroscience (NSF grant HRD 1137725). We also thank Barreto‐Estrada J and Perez‐Acevedo N for her support, collaboration and mentoring during this project.This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

Females have a higher prevalence to develop anxiety disorders, suggesting that estrogen might be involved in female anxiety. Estrogen alters neural activity by affecting neurotransmitter release and the activity of metabotropic glutamate receptors (mGluRs). Whether estradiol interact with group I mGluRs within the basolateral amygdala (BLA) to modulate female anxiety, is unknown. We therefore studied the role of estradiol and group I mGluRs in the BLA, a key region involved in anxiety, by infusing the group I mGluR agonist, (RS)‐3,5‐dihydroxyphenylglycine (DHPG). We used ovariectomized female rats with (OVX‐EB) and without (OVX) estradiol replacement to test whether estradiol‐mGluR interaction modulate female anxiety. We analyzed generalized and anticipatory anxiety using the Elevated Plus Maze and Risk Assessment Behaviors (RABs). In generalized anxiety, DHPG (1.0 μM) increased the percentage time in the open arms, in OVX‐EB but not OVX female rats. In anticipatory anxiety, DHPG (0.1 and 1.0 μM) decreased the number of RABs in OVX‐EB female rats only. DHPG (10.0 μM) did not alter generalized and anticipatory female rat anxiety. We found that: the anxiolytic‐like effect of DHPG depends upon estradiol treatment, suggesting an interaction between estradiol‐mGluR signaling in anxiety. Supported by MBRS‐RISE (GM61838) to DJB, EARDA (NIH‐NIH‐1G11H046326) to PA.

Females are twice affected by anxiety‐related disorders than males. These differences might be due to the effects of gonadal hormones. Metabotropic glutamate receptors (mGluRs) have been implied in the pathophysiology of anxiety. We hypothesized that S‐3,5‐Dihydroxyphenylglycine (DHPG), a group I mGluRs agonist, within the basolateral amygdala (BLA) will modulate anxiety according to sex. Intact males were compared to ovariectomized female rats that contained empty implants (OVX) or implants containing estradiol (OVX‐EB). We used the Vogel Conflict Test (VCT) to analyze conflict‐based anxiety and the Cat Odor Test (COT) for innate anxiety behaviors. In the VCT, intact males displayed more punished licks than OVX female rats (p=0.05). DHPG infusion did not alter any VCT parameter in intact male, OVX and OVX‐EB female rats. In COT, intact males showed lesser number of flat back approached (FBA) and spent more time in contact with the cat odor stimuli than OVX female rats (p= 0.01 & 0.04). DHPG infusion decreased FBA (p=0.02) and stretch attended behaviors in OVX female rats only (p=0.03). In males, DHPG reduced the number of freezing behaviors (p=0.004). Our results indicate sex differences in anxiety‐related behaviors which are task‐dependent. Infusion of DHPG within BLA modulates innate anxiety behaviors in OVX female rats. We concluded that mGluRs might be a pharmacological alternative to modulate anxiety.