G. Biggio

Caffeine-Induced Increases in the Brain and Plasma Concentrations of Neuroactive Steroids in the Rat

The effects of caffeine, a naturally occurring stimulant, on the brain and plasma concentrations of neuroactive steroids were examined in the rat. A single intraperitoneal injection of caffeine induced dose- and time-dependent increases in the concentrations of pregnenolone, progesterone, and 3alpha-hydroxy-5alpha-pregnan-20-one (allopregnanolone) in the cerebral cortex. The increases were significant at a caffeine dose of 25 mg/kg and greatest (+188, +388, and +71%, respectively) at a dose of 100 mg/kg in rats killed 30 min after caffeine administration. Caffeine also increased the plasma concentrations of pregnenolone and progesterone with a dose-response relation similar to that observed in the brain, whereas the caffeine-induced increase in the plasma concentration of allopregnanolone was maximal at a dose of 50 mg/kg. Caffeine increased the plasma concentration of corticosterone, but it had no effect on the brain or plasma concentrations of 3alpha, 21-dihydroxy-5alpha-pregnan-20-one and dehydroepiandrosterone. Moreover, the brain and plasma concentrations of pregnenolone, progesterone, and allopregnanolone were not affected by caffeine in adrenalectomized-orchiectomized rats. These results suggest that neuroactive steroids may modulate the stimulant and anxiogenic effects of caffeine.

Valproic Acid-Hydrophilic Cyclodextrin Complexes and Valproic Acid-Solid Dispersions: Evaluation of Their Potential Pharmaceutical Use

The purpose of this study was to evaluate the potential use of two novel solid formulations of valproic acid (VPA) prepared by complexation with hydrophilic cyclodextrins (CDs) as hydroxypropyl‐β‐ and sulfobutylether‐β‐cyclodextrin and by solid dispersion (SD) in hydrophilic carriers as polyethylene glycol 6000 (PEG 6000) and polyvinylpyrrolidone K‐30 (PVP K‐30). The corresponding cyclodextrin‐based complexes were prepared by the freeze‐drying method while the solid dispersions were obtained by the solvent method. Valproic acid solubility improved by CDs complexation and solid dispersion techniques. Comparison of dissolution profiles with that of VPA sodium salt (NaVP) was made by using release parameters such as dissolution efficiency, percent of drug dissolved after 60 min, and difference and similarity factors. Based on difference and similarity factors, it can be concluded that all the VPA formulations possess dissolution profiles essentially equivalent to those of NaVP at pH 6. However, this conclusion is not confirmed by using the analysis of variance (ANOVA) approach, indicating some significant differences between some SD‐based formulations and NaVP at that pH value. Preliminary pharmacological studies in the pentylenetetrazole test in rats showed some important differences among the SD‐based formulations, NaVP, and VPA as oil/water emulsion. Some implications and limitations of the investigated formulations are discussed.

Age-related changes of benzodiazepine and GABA binding sites in the rat retina.

The changes in the number and sensitivity of benzodiazepine and GABA binding sites in the rat retina during postnatal development, adulthood and ageing and their functional relationship at different ages have been studied. Data indicate an increase in the total number of both GABA and benzodiazepine binding sites with age. In contrast, the activation of retinal benzodiazepine receptor binding by GABA is significantly reduced in aged rats with respect to young adult and newborn rats. Moreover, the activation of retinal benzodiazepine receptor binding induced by dark exposure of the animals is present in young adult rats but is lost in aged rats. These results suggest that in the retina of aged rats there is an increase of GABA and benzodiazepine receptors which have lost their functional connection.

Brain Excitatory/Inhibitory Circuits Cross-Talking with Chromogranin A During Hypertensive and Hibernating States

To date, many scientific attempts have been directed towards the development of experimental models for the identification of neuronal mechanisms evoking cardiovascular and hemodynamic dysfunctions. The spontaneously hypertensive rat (SHR), a genetic model of essential hypertension, has become a valuable rodent for the characterization of molecular markers in hypertensive-related diseases. Recently, growing interests have also been directed to a new experimental paradigm i.e. hibernation, a physiological state consenting the hamster (Mesocricetus auratus) to activate protective mechanisms against ischemic-like complications during the arousal phase. With this intention, the present review will focus attention on specific neurosignaling systems involved with the preservation of hemodynamic conditions in those brain areas that play a pivotal role on such a feature. It is widely known that healthy neurons conserve their structural and responsiveness properties in presence of a constant blood supply, which is assured by their coupling to microvessels and perivascular astrocytes as well as by secretory proteins such as chromogranin A (CgA). So, it will be interesting to establish if this protein alone or with the participation of excitatory/inhibitory neurosignals is capable of influencing some brain areas controlling cardiovascular conditions in both SHRs and hibernating hamsters. In this context, the present work will deliver the most important findings regarding neuronal CgA and its cross-talking ability with major inhibitory (GABA/adenosine) and/or excitatory (glutamate) neuroreceptor systems in relation to hypertensive/hypotensive states of both animal models. Indications deriving from such approaches may provide clinically useful insights regarding their role as protective factors of hemodynamic and neurological disorders.

Molecular mechanisms of tolerance to and withdrawal of GABA(A) receptor modulators

catenin, known to serve as a linker between the cadherin and actin cytoskeleton, results in destabilization of synaptic contacts. On the contrary, overexpression of this catenin causes excess spine formation and reduced spine turnover. Pharmacological suppression of neural activities in hippocampal cultures induces a release of a N-catenin from synapses, whereas elevation of neural activities have opposite effects, i.e., enrichment of this molecule in synapses, suggesting the existence of an activity-dependent mechanism to control the association of a Ncatenin with synapses. In addition, it is known that a number of different cadherin subtypes with distinct adhesive specificities, generated due to the sequence diversity of their extracellular domain, are expressed in the nervous system, and each neuron has a unique set of these cadherins. These observations suggest that the cadherin–catenin complex regulates synapse dynamics from the cytoplasmic side, and possibly synaptic specificity from the extracellular side, although the latter idea is awaiting experimental tests. PL1 CONTROL OF SYNAPTIC JUNCTION DYNAMICS: ROLES OF THE CADHERIN–CATENIN COMPLEX Takeichi, M. RIKEN Center for Developmental Biology, Kobe, Japan

Neurosteroids alter GABA-A receptor subunit gene expression and the action of ethanol on mesocortical dopamine neurons

The progesterone metabolite 5a-pregnan-3a-ol-20-one (5a3a) is a potent allosteric modulator of the GABAA receptor. Some of the pharmacological properties of the GABAA receptor are dependent upon the subunit composition of the receptor. The influence of subunit composition of the human GABAA receptor upon the enhancement of GABA-evoked responses by 5a3a was examined using the Xenopus laevis oocyte expression system and the two electrode voltage-clamp technique. Steroid potency was modestly affected by the aor c-isoform present within the ternary complex. The b-isoform did not influence the GABAmodulatory effect of the steroid. Incorporation of the e-subunit suppressed, whereas the presence of the d-subunit enhanced the actions of 5a3a. To determine the effects of 5a3a on GABA-ergic synaptic transmission the patch clamp technique was applied to rat hippocampal slices. Bicuculline-sensitive miniature inhibitory postsynaptic currents (mIPSCs) were recorded from CA1 pyramidal and dentate granule cells. In CA1 neurones physiological concentrations of 5a3a (30– 100 nM) significantly prolonged mIPSCs decay whereas dentate gyrus granule (DG) neurones were relatively insensitive to the steroid. By contrast, the metabolically stable analogue of 5a3a, ganaxolone (3b-methyl-3a-ol-5a-pregnan-20-one) induced a significant prolongation of mIPSCs recorded from DG neurones at concentrations which are equi-effective with 5a3a at recombinant GABAA receptors (i.e. 30 and 100 nM). These results suggest that neurosteroid modulation of GABAA receptor-mediated inhibition maybe influenced not only by the subunit composition of the receptor, but also by local steroid metabolism. Acknowledgments: Supported by the MRC (DB).