Organization of Circadian Behavior Relies on Glycinergic Transmission.

The small ventral lateral neurons (sLNvs) constitute a central circadian pacemaker in the Drosophila brain. They organize daily locomotor activity, partly through the release of the neuropeptide pigment-dispersing factor (PDF), coordinating the action of the remaining clusters required for network synchronization. Despite extensive efforts, the basic principles underlying communication among circadian clusters remain obscure. We identified classical neurotransmitters released by sLNvs through disruption of specific transporters. Adult-specific RNAi-mediated downregulation of the glycine transporter or impairment of glycine synthesis in LNv neurons increased period length by nearly an hour without affecting rhythmicity of locomotor activity. Electrophysiological recordings showed that glycine reduces spiking frequency in circadian neurons. Interestingly, downregulation of glycine receptor subunits in specific sLNv targets impaired rhythmicity, revealing involvement of glycine in information processing within the network. These data identify glycinergic inhibition of specific targets as a cue that contributes to the synchronization of the circadian network.

Inferring Evolution of Habitat Usage and Body Size in Endangered, Seasonal Cynopoeciline Killifishes from the South American Atlantic Forest through an Integrative Approach (Cyprinodontiformes: Rivulidae).

Cynopoecilines comprise a diversified clade of small killifishes occurring in the Atlantic Forest, one of the most endangered biodiversity hotspots in the world. They are found in temporary pools of savannah-like and dense forest habitats, and most of them are highly threatened with extinction if not already extinct. The greatest gap in our knowledge of cynopoecilines stems from the absence of an integrative approach incorporating molecular phylogenetic data of species still found in their habitats with phylogenetic data taken from the rare and possibly extinct species without accessible molecular information. An integrative analysis combining 115 morphological characters with a multigene dataset of 2,108 bp comprising three nuclear loci (GLYT1, ENC1, Rho), provided a robust phylogeny of cynopoeciline killifishes, which was herein used to attain an accurate phylogenetic placement of nearly extinct species. The analysis indicates that the most recent common ancestor of the Cynopoecilini lived in open vegetation habitats of the Atlantic Forest of eastern Brazil and was a miniature species, reaching between 25 and 28 mm of standard length. The rare cases of cynopoecilines specialized in inhabiting pools within dense forests are interpreted as derived from four independent evolutionary events. Shifts in habitat usage and biogeographic patterns are tentatively associated to Cenozoic paleogeographic events, but the evolutionary history of cynopoecilines may be partially lost by a combination of poor past sampling and recent habitat decline. A sharp evolutionary shift directed to increased body size in a clade encompassing the genera Campellolebias and Cynopoecilus may be related to a parallel acquisition of an internally-fertilizing reproductive strategy, unique among aplocheiloid killifishes. This study reinforces the importance of adding morphological information to molecular databases as a tool to understand the biological complexity of organisms under intense pressure from loss of habitat.

Low expression of Gria1 and Grin1 glutamate receptors in the nucleus accumbens of Spontaneously Hypertensive Rats (SHR).

The Spontaneously Hypertensive Rat (SHR) strain is a classical animal model for the study of essential hypertension. Recently, our group suggested that this strain could be a useful animal model for schizophrenia, which is a severe mental illness with involvement of glutamatergic system. The aim of this study is to investigate glutamatergic receptors (Gria1 and Grin1) and glycine transporter (Glyt1) gene expression in the prefrontal cortex (PFC) and nucleus accumbens (NAcc) of SHR animals. The effects in gene expression of a chronic treatment with antipsychotic drugs (risperidone, haloperidol and clozapine) were also analyzed. Animals were treated daily for 30 days, and euthanized for brain tissue collection. The expression pattern was evaluated by Real Time Reverse-Transcriptase (RT) PCR technique. In comparison to control rats, SHR animals present a lower expression of both NMDA (Grin1) and AMPA (Gria1) gene receptors in the NAcc. Antipsychotic treatments were not able to change gene expressions in any of the regions evaluated. These findings provide evidence for the role of glutamatergic changes in schizophrenia-like phenotype of the SHR strain.

Glycine transporters (glycine transporter 1 and glycine transporter 2) are expressed in retina.

The amino acid glycine is an inhibitory neurotransmitter in the spinal cord, brain stem, and vertebrate retina. The effective synaptic concentrations of glycine are regulated by at least two transporters: glycine transporter 1 and glycine transporter 2. Here, we show retinal expression of glycine transporter 1 by in-situ hybridization and of glycine transporter 2 by reverse transcriptase-PCR and in-situ hybridization. In-situ hybridization signals were observed in the ganglionar and inner nuclear layer as well as in the outer nuclear layer of the frog and rat retinas. In addition, accumulation of H-glycine was observed in isolated photoreceptor cells. The expression of these transporters in nonglycinergic cells suggests that they may also modulate electrical signals.

Pharmacological properties of glycine uptake in the developing rat retina.

A pharmacological characterization of glycine transport was performed in the rat retina at different postnatal ages. The uptake of 3H-glycine increased during the first 2 weeks of postnatal age, reaching maximum values at 12 days; then it decreased sharply to the adult values. We found a Na+ -dependent and high-affinity transport system with a Km of 100 microM. The Na+ Hill coefficient for glycine uptake was 1.76 +/- 0.07. Although glycine uptake was insensitive to staurosporine and phorbol ester, it was reduced 40-50% by sarcosine and ALX5407. Besides, amoxapine inhibited glycine uptake by 40 and 70% in adult and immature retina, respectively. These results suggest that the Glyt1 transporter was concentrated in the nerve terminals. In addition to the presence of Glyt1 in the retina, our results provided evidence of the occurrence of Glyt2 and/or another isoform of glycine transporter, which might have had a role in the retina development.