Paolo N. Catalano

GABA B1 Knockout Mice Reveal Alterations in Prolactin Levels, Gonadotropic Axis, and Reproductive Function

γ-Aminobutyric acid (GABA) has been implicated in the control of hypophyseal functions. We evaluated whether the constitutive loss of functional GABAB receptors in GABAB1 knockout (GABAB1–/–) mice alters hormonal levels, under basal and stimulated conditions, and reproductive function. The serum hormone levels were measured by radioimmunoassay, the estrous cyclicity was evaluated by vaginal lavages, and the mating behavior was determined by the presence of vaginal plugs. A moderate hyperprolactinemic condition was observed, in which prolactin increase and thyroid-stimulating hormone decrease were similar between genotypes. Basal luteinizing hormone (LH), follicle-stimulating hormone, thyroid-stimulating hormone, and growth hormone levels were similar between genotypes in each sex. Analysis of the gonadotropin axis revealed no differences in puberty onset between female genotypes. In con trast, the estrous cyclicity was significantly disrupted in GABAB1–/– female mice, showing significantly extended periods in estrus and shortened periods in proestrus. Reproduction was significantly compromised in GABAB1–/– females, with a significantly lower proportion of mice (37.5%) getting pregnant during the first 30 days of mating as compared with wild-type controls (87.5%). Moreover, only 14% of vaginal plug positive GABAB1–/– females had successful pregnancies as compared with 75% in the controls. In addition, the postovariectomy LH rise was significantly advanced in GABAB1–/– mice, while the response to estradiol feedback was similar in both genotypes. In conclusion, our endocrine analysis of GABAB1–/– mice reveals that GABAB receptors are involved in the regulation of basal prolactin titers. Moreover, the hypothalamic-hypophyseal-ovarian axis is seriously disturbed, with alterations in cyclicity, postcastration LH increase, and fertility indexes. The molecular mechanism underlying these hormonal disturbances remains to be addressed.

Lack of functional GABAB receptors alters GnRH physiology and sexual dimorphic expression of GnRH and GAD-67 in the brain

GABA, the main inhibitory neurotransmitter, acts through GABA(A/C) and GABA(B) receptors (GABA(B)Rs); it is critical for gonadotropin regulation. We studied whether the lack of functional GABA(B)Rs in GABA(B1) knockout (GABA(B1)KO) mice affected the gonadotropin axis physiology. Adult male and female GABA(B1)KO and wild-type (WT) mice were killed to collect blood and tissue samples. Gonadotropin-releasing hormone (GnRH) content in whole hypothalami (HT), olfactory bulbs (OB), and frontoparietal cortexes (CT) were determined (RIA). GnRH expression by quantitative real-time PCR (qRT-PCR) was evaluated in preoptic area-anterior hypothalamus (POA-AH), medial basal-posterior hypothalamus (MBH-PH), OB, and CT. Pulsatile GnRH secretion from hypothalamic explants was measured by RIA. GABA, glutamate, and taurine contents in HT and CT were determined by HPLC. Glutamic acid decarboxylase-67 (GAD-67) mRNA was measured by qRT-PCR in POA-AH, MBH-PH, and CT. Gonadotropin content, serum levels, and secretion from adenohypophyseal cell cultures (ACC) were measured by RIA. GnRH mRNA expression was increased in POA-AH of WT males compared with females; this pattern of expression was inversed in GABA(B1)KO mice. MBH-PH, OB, and CT did not follow this pattern. In GABA(B1)KO females, GnRH pulse frequency was increased and GABA and glutamate contents were augmented. POA-AH GAD-67 mRNA showed the same expression pattern as GnRH mRNA in this area. Gonadotropin pituitary contents and serum levels showed no differences between genotypes. Increased basal LH secretion and decreased GnRH-stimulated gonadotropin response were observed in GABA(B1)KO female ACCs. These results support the hypothesis that the absence of functional GABA(B)Rs alters GnRH physiology and critically affects sexual dimorphic expression of GnRH and GAD-67 in POA-AH.

Recent patents on the synthesis and application of silica nanoparticles for drug delivery.

Drug delivery systems are designed to improve therapy efficacy as well as patient compliance. This could be accomplished by specifically targeting a medication intact to its active site, therefore reducing side-effects and enabling high local drug concentrations. Silica nanoparticles have gained ground in the biomedical field for their biocompatibility and biodegradability, being themselves inert and stable, thus enabling a variety of formulation designs for application in the pharmaceutical industry. This paper is a review of the recent patents on the applications of silica nanoparticles for drug delivery and their preparation. The review will focus on the different techniques available to obtain silica nanoparticles with variable morphology and their drug targeting applications, providing an overview of silica particles synthesis described in the literature.

Effect of Androgens on Sexual Differentiation of Pituitary Gamma-Aminobutyric Acid Receptor Subunit GABAB Expression

Previous work demonstrated a sexually dimorphic ontogenic expression of γ-aminobutyric acid receptors (GABABR) in rat pituitary. As sex steroids determine sex-specific expression patterns, we now studied the effect of sex hormones on pituitary GABABR expression. GABABR subunits, measured by Western blot and by semi-quantitative RT-PCR and luteinizing hormone (LH), follicle-stimulating hormone (FSH) and testosterone measured by RIA were determined in two experimental designs: First experimental design: 8- and 15-day-old females (8F, 15F); 8F and 15F treated with 100 µg testosterone propionate (TP) on day 1 of life (8F100TP, 15F100TP), 8- and 15-day-old males (8M, 15M) and 8M and 15M castrated on day 1 (8MC, 15MC). Second experimental design: 8-day-old female and male animals: 8F, 8F100TP, 8F treated with 1 µg/day TP on days 1–4 (8F1TP), 8F treated with the androgen antagonist Flutamide (Flut: 2.5 mg/100 g BW of pregnant mother on days E17-E23) (8F-Flut), 8M, 8MC, 8M treated with Flut as above (8M-Flut) and 8MC-Flut. In these animals, in addition, GABA, glutamate, aspartate and taurine were measured by HPLC in hypothalami and cortex. In the first set of experiments, GABAB1R mRNA/protein expression was higher in 8F than in 15F, 8M or 15M. In 8F100TP, GABAB1R mRNA/protein decreased to male levels. TP treatment did not alter GABAB1R expression in 15F. There was no difference in GABAB1R expression between 8M and 15M and neonatal castration did not modify its expression. In the second set of experiments, TP (1 µg) or Flut did not modify GABAB1R in 8F, while 100 µg TP continued to decrease GABAB1R expression. In 8M, Flut, alone or with castration, increased GABAB1R mRNA/protein expression to 8F. Hypothalamic GABA content followed the same pattern as pituitary GABABR expression in 8-day-old animals, suggesting a cross-regulation. With regard to hormonal levels, 100 µg, but not 1 µg TP altered gonadotropins at 8 days, although both treatments effectively androgenized females as evidenced by lack of cycling. We conclude that androgens, acting pre- and postnatally, decrease pituitary GABABR subunit expression.

Lack of functional GABAB receptors alters Kiss1 , Gnrh1 and Gad1 mRNA expression in the medial basal hypothalamus at postnatal day 4.

Background/Aims: Adult mice lacking functional GABAB receptors (GABAB1KO) show altered Gnrh1 and Gad1 expressions in the preoptic area-anterior hypothalamus (POA-AH) and females display disruption of cyclicity and fertility. Here we addressed whether sexual differentiation of the brain and the proper wiring of the GnRH and kisspeptin systems were already disturbed in postnatal day 4 (PND4) GABAB1KO mice. Methods: PND4 wild-type (WT) and GABAB1KO mice of both sexes were sacrificed; tissues were collected to determine mRNA expression (qPCR), amino acids (HPLC), and hormones (RIA and/or IHC). Results: GnRH neuron number (IHC) did not differ among groups in olfactory bulbs or OVLT-POA. Gnrh1 mRNA (qPCR) in POA-AH was similar among groups. Gnrh1 mRNA in medial basal hypothalamus (MBH) was similar in WTs but was increased in GABAB1KO females compared to GABAB1KO males. Hypothalamic GnRH (RIA) was sexually different in WTs (males > females), but this sex difference was lost in GABAB1KOs; the same pattern was observed when analyzing only the MBH, but not in the POA-AH. Arcuate nucleus Kiss1 mRNA (micropunch-qPCR) was higher in WT females than in WT males and GABAB1KO females. Gad1 mRNA in MBH was increased in GABAB1KO females compared to GABAB1KO males. Serum LH and gonadal estradiol content were also increased in GABAB1KOs. Conclusion: We demonstrate that GABABRs participate in the sexual differentiation of the ARC/MBH, because sex differences in several reproductive genes, such as Gad1, Kiss1 and Gnrh1, are critically disturbed in GABAB1KO mice at PND4, probably altering the organization and development of neural circuits governing the reproductive axis.

GABAB Receptors in Neuroendocrine Regulation

Gamma-amino butyric acid (GABA), in addition to being a metabolic intermediate and the main inhibitory neurotransmitter in the synaptic cleft, is postulated as a neurohormone, a paracrine signaling molecule, and a trophic factor. It acts through pre- and post-synaptic receptors, named GABAA and GABAC (ionotropic receptors) and GABAB (metabotropic receptor). Here we reviewed the participation of GABAB receptors in the regulation of the hypothalamic-pituitary-gonadal axis, using physiological, biochemical, and pharmacological approaches in rats, as well as in GABAB1 knock-out mice, that lack functional GABAB receptors. Our general conclusion indicates that GABAB receptors participate in the regulation of pituitary hormone secretion acting both in the central nervous system and directly on the gland. PRL and gonadotropin axes are affected by GABAB receptor activation, as demonstrated in the rat and also in the GABAB1 knock-out mouse. In addition, hypothalamic and pituitary GABAB receptor expression is modulated by steroid hormones. GABA participation in the brain control of pituitary secretion through GABAB receptors depends on physiological conditions, being age and sex critical factors.These results indicate that patients receiving GABAB agonists/antagonists should be monitored for possible endocrine side effects.

Surface chemistry of nanobiomaterials with antimicrobial activity

Abstract Biofilms are common in nature and are often associated with undesirable effects such as the deterioration of concrete, metal corrosion, fouling of oil and gas pipelines and ships hulls, dental plaques causing tooth decay, and microbial colonization of indwelling or percutaneous medical devices, such as catheters, artificial valves, joints, and stents. Nanomaterials with distinctive physicochemical properties and high surface areas are offering new alternatives for the development of antibiofilm and bactericidal surfaces. In order to obtain antimicrobial coatings several approaches will be described. One approach consists of using a controlled-release nanostructured coating, in which the antibiotic is released from the biomedical device and intercepts bacteria in the vicinity. The disadvantage of the release approach is that the duration and effectiveness of antibacterial action are limited by loading and release kinetics. The second approach consists on the application of a molecular surface layer of covalently immobilized (“grafted”) molecules or on the modification of surface nanotopography, which can prevent bacterial attachment to material surfaces by either killing bacteria or changing physicochemical characteristics of the surfaces (hydrophobicity/hydrophilicity, charge, surface free energy, nanoroughness). In addition, much longer, perhaps indefinite, effectiveness can be ascertained by using this approach.

Innovative Immobilization Matrices.

We present a brief survey of some of the recent work of Professor Luis E. Díaz, performed together with his students and collaborators at the University of Buenos Aires. Dr Luis E. Díaz has been involved in research on biochemical and pharmaceutical sciences solving scientific and industry problems for over 40 years until he passed away. Prof. Díaz scientific interests included various topics from NMR spectroscopy to biomedicine but fundamentally he focused in various aspects of chemistry (analytical, organic, inorganic and environmental). This is not a complete survey but a sampling of prominent projects related to sol-gel chemistry with a focus on some of his recent publications.