Pyramidal neurons in the medial prefrontal cortex (mPFC) critically contribute to cocaine-seeking behavior in humans and rodents. Activity of these neurons is significantly modulated by GABAergic, parvalbumin-containing, fast-spiking interneurons, the majority of which are enveloped by specialized structures of extracellular matrix called perineuronal nets (PNNs), which are integral to the maintenance of many types of plasticity. Using a conditioned place preference (CPP) procedure, we found that removal of PNNs primarily from the prelimbic region of the mPFC of adult, male, Sprague Dawley rats impaired the acquisition and reconsolidation of a cocaine-induced CPP memory. This impairment was accompanied by a decrease in the number of c-Fos-positive cells surrounded by PNNs. Following removal of PNNs, the frequency of inhibitory currents in mPFC pyramidal neurons was decreased; but following cocaine-induced CPP, both frequency and amplitude of inhibitory currents were decreased. Our findings suggest that cocaine-induced plasticity is impaired by removal of prelimbic mPFC PNNs and that PNNs may be a therapeutic target for disruption of cocaine CPP memories.

Neurons in the upper lumbar spinal cord project axons containing gastrin-releasing peptide (GRP) to innervate lower lumbar regions controlling erection and ejaculation. This system is vestigial in female rats and in males with genetic dysfunction of androgen receptors, but in male rats, pharmacological stimulation of spinal GRP receptors restores penile reflexes and ejaculation after castration. GRP offers new avenues for understanding potential therapeutic approaches to masculine reproductive dysfunction.GRP, a member of the bombesin-like peptide family 1 , is distributed widely in the central nervous system and gastrointestinal tract of mammals 2 , 3 . GRP and neuromedin B (NMB), the mammalian counterpart of bombesin, play a role in many physiological processes, including itch 4 , circadian rhythms 5 , food intake 6 and fear memory consolidation 7 , 8 . In mammals, bombesin-like peptides act through a family of at least three G protein-coupled receptors: GRP-preferring receptor (GRP-R), NMB-preferring receptor (NMB-R) and bombesin receptor subtype-3 (BRS-3) 9 .Using immunocytochemistry (ICC) directed at GRP, we found a group of neurons within a region previously dubbed the 'spinal ejaculation generator' because toxins that selectively lesion galanin-containing neurons there virtually eliminate ejaculation in rats 10 . These galaninergic neurons project to the thalamus 10 , but it had been unclear whether there are also direct connections between this center and the lower spinal cord regions that directly trigger ejaculation 11 . The separate, GRP-containing neurons that we found within the center projected axons to more caudal spinal regions and were much more prominent in wild-type (WT) males than in WT females ( Fig. 1a,b; Supplementary Fig. 1 online) (n = 5, F 2,12 = 299.9, P < 0.001). Semiquantitative reverse transcription (RT)-PCR confirmed more pre-pro NIH-PA Author ManuscriptNIH-PA Author Manuscript NIH-PA Author ManuscriptGrp transcripts in this region of males than of females ( Supplementary Fig. 2 online). To test whether androgen receptors direct sexual dimorphism of these neurons, we examined genetically male (XY) Long-Evans rats carrying the testicular feminization mutation (Tfm) of the androgen receptor gene Ar. These rats develop testes embryologically and secrete testosterone pre-and postnatally but, because their androgen receptor protein is dysfunctional, develop a wholly feminine exterior, including a clitoris rather than a penis. The spinal cord of Tfm rats was hyperfeminine, having even fewer GRP-positive neurons in this region than did WT females (P <0.001) (Fig. 1c,d). In normal males, GRP-expressing neurons also expressed androgen receptor (96.1 ± 1.7%; n = 4 WT males), but not estrogen receptor alpha (ERα) (Fig. 1e-h). Because androgens such as testosterone augment ejaculation in male rats and humans 12 , the presence of androgen receptor in the GRPpositive neurons of the ejaculation center offers a locus for androgenic modulation of ejaculation and other sexual reflexes.ICC r...

Many studies demonstrate that exposure to testicular steroids such as testosterone early in life masculinizes the developing brain, leading to permanent changes in behavior. Traditionally, masculinization of the rodent brain is believed to depend on estrogen receptors (ERs) and not androgen receptors (ARs). According to the aromatization hypothesis, circulating testosterone from the testes is converted locally in the brain by aromatase to estrogens, which then activate ERs to masculinize the brain. However, an emerging body of evidence indicates that the aromatization hypothesis cannot fully account for sex differences in brain morphology and behavior, and that androgens acting on ARs also play a role. The testicular feminization mutation (Tfm) in rodents, which produces a nonfunctional AR protein, provides an excellent model to probe the role of ARs in the development of brain and behavior. Tfm rodent models indicate that ARs are normally involved in the masculinization of many sexually dimorphic brain regions and a variety of behaviors, including sexual behaviors, stress response and cognitive processing. We review the role of ARs in the development of the brain and behavior, with an emphasis on what has been learned from Tfm rodents as well as from related mutations in humans causing complete androgen insensitivity. Keywordscomplete androgen insensitivity syndrome -CAIS; vasopressin; anxiety; spatial memory; bed nucleus; medial amygdala; SDN-POA; sexual behavior; aggression; VMH Exposure to testicular steroids such as testosterone (T) early in life masculinizes the developing brain, leading to permanent changes in behavior in a wide variety of animal models (Morris et al., 2004). According to the aromatization hypothesis, T is converted by aromatase into 17-β estradiol (E2), which then acts on estrogen receptors (ERs) to masculinize the brain (Naftolin et al., 1975). Traditionally, aromatization is believed to be the mechanism by which the rodent brain becomes masculinized and defeminized. Some sexually dimorphic regions within the hypothalamus adhere well to this hypothesis, including the sexually dimorphic nucleus of the preoptic area (SDN-POA) and anteroventral periventricular nucleus (AVPV). T spares neuronsCorresponding author: Damian Zuloaga, Neuroscience Program, 108 Giltner Hall, Michigan State University, East Lansing, MI 48824, Phone: Fax: (517) 432-2744, Email: E-mail: zuloagad@msu.edu. Publisher's Disclaimer: This is a PDF file of an unedited manuscript that has been accepted for publication. As a service to our customers we are providing this early version of the manuscript. The manuscript will undergo copyediting, typesetting, and review of the resulting proof before it is published in its final citable form. Please note that during the production process errors may be discovered which could affect the content, and all legal disclaimers that apply to the journal pertain. NIH Public Access NIH-PA Author ManuscriptNIH-PA Author Manuscript NIH-PA Author Manuscript from death in the SDN-POA...

Estrogen receptors regulate multiple brain functions including stress, sexual, and memory associated behaviors as well as control of neuroendocrine and autonomic function. During development, estrogen signaling is involved in programming adult sex differences in physiology and behavior. Expression of estrogen receptor alpha changes across development in a region specific fashion. By contrast, estrogen receptor beta (ERβ) is expressed in many brain regions, yet few studies have explored sex and developmental differences in its expression largely due to the absence of selective reagents for anatomical localization of the protein. In this study, we utilized bacterial artificial chromosome transgenic mice expressing ERβ identified by enhanced green fluorescent protein (EGFP) to compare expression levels and distribution of ERβ in the male and female mouse forebrain on the day of birth (P0), postnatal day 4 (P4) and P21. Using qualitative analysis, we mapped the distribution of ERβ–EGFP and found developmental alterations in ERβ expression within the cortex, hippocampus, and hypothalamic regions including the arcuate, ventromedial, and paraventricular nuclei. We also report a sex difference in ERβ in the bed nucleus of the stria terminalis with males showing greater expression at P4 and P21. Another sex difference was found in the anteroventral periventricular nucleus of P21, but not P0 or P4 mice, where ERβ-EGFP-ir cells were densely clustered near the 3rd ventricle in females but not males. These developmental changes and sex differences in ERβ indicate a mechanism through which estrogens may differentially affect brain functions or program adult physiology at select times during development.

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