Endogenous neurosteroids have rapid actions on ion channels, particularly GABA(A) receptors, which are potentiated by nanomolar concentrations of 3alpha-hydroxypregnane neurosteroids. Previous evidence suggests that 3beta-hydroxypregnane steroids may competitively antagonize potentiation induced by their 3alpha diastereomers. Because of the potential importance of antagonists as experimental and clinical tools, we characterized the functional effect of 3beta-hydroxysteroids. Although 3beta-hydroxysteroids reduced the potentiation induced by 3alpha-hydroxysteroids, 3beta-hydroxysteroids acted noncompetitively with respect to potentiating steroids and inhibited the largest degrees of potentiation most effectively. Potentiation by high concentrations of barbiturates was also reduced by 3beta-hydroxysteroids. 3beta-Hydroxysteroids are also direct, noncompetitive GABA(A) receptor antagonists. 3beta-Hydroxysteroids coapplied with GABA significantly inhibited responses to > or =15 microm GABA. The profile of block was similar to that exhibited by sulfated steroids, known blockers of GABA(A) receptors. This direct, noncompetitive effect of 3beta-hydroxysteroids was sufficient to account for the apparent antagonism of potentiating steroids. Mutated receptors exhibiting decreased sensitivity to sulfated steroid block were insensitive to both the direct effects of 3beta-hydroxysteroids on GABA(A) responses and the reduction of potentiating steroid effects. At concentrations that had little effect on GABAergic synaptic currents, 3beta-hydroxysteroids and low concentrations of sulfated steroids significantly reversed the potentiation of synaptic currents induced by 3alpha-hydroxysteroids. We conclude that 3beta-hydroxypregnane steroids are not direct antagonists of potentiating steroids but rather are noncompetitive, likely state-dependent, blockers of GABA(A) receptors. Nevertheless, these steroids may be useful functional blockers of potentiating steroids when used at concentrations that do not affect baseline neurotransmission.

Sulfated steroids like pregnenolone sulfate (PS) are found endogenously in the central nervous system where they may modulate GABAA receptors. Understanding the mechanism of steroid inhibition is important for understanding the conditions under which endogenous steroids modulate GABAA receptor function, assessing their potential clinical utility, and for evaluating sulfated steroids as probes of receptor behaviour. Some previous studies suggest that sulfated steroid inhibition exhibits activation dependence, whilst other studies suggest only slow, time‐dependent inhibition, perhaps reflecting slow PS association with receptors. We tested activation dependence in several ways. Steroid potency increased 2‐ to 3‐fold with ≈10‐fold change in GABA concentration. PS inhibition of saturating partial agonist responses suggested that the level of channel activation, rather than receptor occupancy by agonist, is important for PS inhibition. Inhibition by sulfated steroids exhibited weak or no voltage dependence. Responses to rapid applications of exogenous GABA differed little whether PS was pre‐applied or simply co‐applied with GABA, consistent with the hypothesis that the actions of PS are facilitated by receptor activation. PS applied during steady‐state GABA responses exhibited slow onset and offset rate constants. The offset, rather than onset, was significantly slowed by elevated GABA concentration. At hippocampal synapses, large, multiquantal IPSCs were inhibited more effectively by a fixed concentration of PS than small quantal content IPSCs, consistent with known ‘pooling’ of transmitter following multiquantal release. Picrotoxinin, although superficially similar to PS in its activation dependence, was dissimilar from PS in a number of details. In summary, PS inhibition exhibits activation dependence that may be explained by activation‐dependent binding and altered desensitization.

Dietary emulsifiers carboxylmethylcellulose (CMC) and polysorbate 80 (P80) alter the composition of the intestinal microbiota and induce chronic low-grade inflammation, ultimately leading to metabolic dysregulations in mice. As both gut microbiota and intestinal health can influence social and anxiety-like behaviors, we investigated whether emulsifier consumption would detrimentally influence behavior. We confirmed that emulsifier exposure induced chronic intestinal inflammation, increased adiposity, and altered gut microbiota composition in both male and female mice, although the specific microboal taxa altered following emulsifier consumption occurred in a sex-dependent manner. Importantly, emulsifier treatment altered anxiety-like behaviors in males and reduced social behavior in females. It also changed expression of neuropeptides implicated in the modulation of feeding as well as social and anxiety-related behaviors. Multivariate analyses revealed that CMC and P80 produced distinct clustering of physiological, neural, and behavioral effects in male and female mice, suggesting that emulsifier treatment leads to a syndrome of sex-dependent changes in microbiota, physiology, and behavior. This study reveals that these commonly used food additives may potentially negatively impact anxiety-related and social behaviors and may do so via different mechanisms in males and females.

BackgroundGut dysbiosis is observed in several neuropsychiatric disorders exhibiting increases in anxiety behavior, and recent work suggests links between gut inflammation and such disorders. One source of this inflammation may be lipopolysaccharide (LPS), a toxic component of gram-negative bacteria. Here, we (1) determine whether oral gavage of LPS, as a model of gut-derived endotoxemia, affects anxiety-like and/or repetitive behaviors; (2) test whether these changes depend on TLR4 signaling; and (3) test the extent to which gut-derived endotoxin and TLR4 antagonism affects males and females differently.MethodsIn experiment 1, male wild-type (WT) and Tlr4−/− mice were tested for locomotor, anxiety-like, and repetitive behaviors in an automated open field test apparatus, 2 h after oral gavage of LPS or saline. In experiment 2, male and female WT mice received an oral gavage of LPS and an injection of one or two TLR4 antagonists that target different TLR4 signaling pathways ((+)-naloxone and LPS derived from R. sphaeroides (LPS-RS)). Univariate and multivariate analyses were used to identify effects of treatment, sex, and genotype and their interaction.ResultsIn experiment 1, oral gavage of LPS increased anxiety-like behavior in male WT mice but not in Tlr4−/− mice. In experiment 2, oral gavage of LPS increased anxiety-like and decreased repetitive behaviors in WT mice of both sexes. Neither antagonist directly blocked the effects of orally administered LPS. However, treatment with (+)-naloxone, which blocks the TRIF pathway of TLR4, had opposing behavioral effects in males and females (independent of LPS treatment). We also identified sex differences in the expression of interleukin-6, a pro-inflammatory cytokine, in the gut both in basal conditions and in response to LPS.ConclusionIn spite of the ubiquitous nature of LPS in the gut lumen, this is the first study to demonstrate that intestinally derived LPS can initiate behavioral aspects of the sickness response. While an increased enteric load of LPS increases anxiety-like behavior in both sexes, it likely does so via sex-specific mechanisms. Similarly, TLR4 signaling may promote baseline expression of repetitive behavior differently in males and females. This study lays the groundwork for future interrogations into connections between gut-derived endotoxin and behavioral pathology in males and females.Electronic supplementary materialThe online version of this article (10.1186/s13293-018-0166-x) contains supplementary material, which is available to authorized users.

Brattleboro rats harbor a spontaneous deletion of the arginine-vasopressin (Avp) gene. In addition to diabetes insipidus, these rats exhibit low levels of anxiety and depressive behaviors. Recent work on the gut-brain axis has revealed that gut microbiota can influence anxiety behaviors. Therefore, we studied the effects of Avp gene deletion on gut microbiota. Since Avp gene expression is sexually different, we also examined how Avp deletion affects sex differences in gut microbiota. Males and females show modest but differentiated shifts in taxa abundance across 3 separate Avp deletion genotypes: wildtype (WT), heterozygous (Het) and AVP-deficient Brattleboro (KO) rats. For each sex, we found examples of taxa that have been shown to modulate anxiety behavior, in a manner that correlates with anxiety behavior observed in homozygous knockout Brattleboro rats. One prominent example is Lactobacillus, which has been reported to be anxiolytic: Lactobacillus was found to increase in abundance in inverse proportion to increasing gene dosage (most abundant in KO rats). This genotype effect of Lactobacillus abundance was not found when females were analyzed independently. Therefore, Avp deletion appears to affect microbiota composition in a sexually differentiated manner.