High production of poly(3-hydroxybutyrate) from a wild Bacillus megaterium Bolivian strain.

AIM: Taking into account that a novel strain of Bacillus megaterium was isolated from Uyuni salt lake (Bolivia) in a previous work, the objectives of this new study were to determine the maximal Poly-3-hydroxybutyrate production potential of B. megaterium strain uyuni S29 in an industrial conventional media, the possibility that the strain accumulates different types of polyhydroxyalkanoates, the cellular morphology during the biosynthesis process and the characterization of the produced biopolymers. METHODS AND RESULTS: The micro-organism was first tested in a 3-L bioreactor obtaining a high specific growth rate of 1·64 h(-1). A second fed-batch experiment was carried out in shaking flasks, reaching up to 70% PHB of cell dry mass. The biosynthesized polymers were extracted by two different extraction procedures and characterized. The results showed that all of them were PHB with thermal properties different to the conventional PHB. The micrographs taken by TEM show the different cell morphology during the fermentation process. CONCLUSIONS: In this previous study, the strain not only grew properly in the industrial conditions proposed without spore formation, but also produced and accumulated a large content of PHB, never reached before for its genus. Therefore, if the culture conditions can be optimized, the biopolymer production could be increased. SIGNIFICANCE AND IMPACT OF THE STUDY: The impact of the study has related to the area of the biomaterials and their production. The study provides new data related to the high production of PHB from the wild novel strain B. megaterium uyuni S29, the highest polymer accumulation for the genus Bacillus without spores formation.

Spontaneous activity in the developing gerbil auditory cortex in vivo involves GABAergic transmission.

A salient feature of the developing brain is that spontaneous oscillations (SOs) and waves may influence the emergence of synaptic connections. While GABA produces depolarization and may support SOs in the neurons of developing rodents, it elicits hyperpolarization and diminishes SOs in developing gerbil auditory cortex (ACx). Therefore, we asked whether SOs exist in developing gerbil ACx in vivo and if GABAergic involvement can be manipulated. In vivo extracellular recordings in P3-5 ACx revealed SOs with longer burst durations and shorter inter-event intervals compared to ACx SOs in slices. ACx was then validated by gross anatomical features and lesions created at the in vivo recording site that corresponded with the electrophysiological coordinates of thalamorecipient ACx in slices. Further, NeuroVue Red, a lipophilic dye loaded at the in vivo recording sites, stained anatomically identifiable fiber tracks between the ACx and the auditory thalamus, medial geniculate body (MG). Separately, to chronically perturb GABAergic role in SOs, P2-5 pups were administered daily with GABA(A) receptor blocker, bicuculline (BIC). We then recorded from P14-17 ACx neurons in slices generated after hearing onset. ACx neurons from BIC-administered pups exhibited spontaneous action potentials in contrast to subthreshold synaptic potentials in neurons from sham-injected animals. Finally, to elucidate whether the gap junction blocker mefloquine (MFQ) previously shown to dampen ACx SOs in slices affected GABAergic transmission, MFQ was acutely applied in P3-5 slices while spontaneous inhibitory postsynaptic currents (sIPSCs) were recorded. Whereas MFQ increased the amplitude and frequency of sIPSCs in ACx neurons, the broad-spectrum gap junction blocker carbenoxolone decreased sIPSC amplitudes only. Together, we show that P2-5 gerbil ACx can endogenously generate SOs in vivo. Persistence of activity in ACx in P14-17 slices from pups administered with BIC at P2-5 implies that inhibitory GABAergic activity linked with gap-junction participates in the maturation of ACx.

Direct measurement of single-channel Ca(2+) currents in bullfrog hair cells reveals two distinct channel subtypes.

1. To confer their acute sensitivity to mechanical stimuli, hair cells employ Ca(2+) ions to mediate sharp electrical tuning and neurotransmitter release. We examined the diversity and properties of voltage-gated Ca(2+) channels in bullfrog saccular hair cells by means of perforated and cell-attached patch-clamp techniques. Whole-cell Ca(2+) current records provided hints that hair cells express L-type as well as dihydropyridine-insensitive Ca(2+) currents. 2. Single Ca(2+) channel records confirmed the presence of L-type channels, and a distinct Ca(2+) channel, which has sensitivity towards omega-conotoxin GVIA. Despite its sensitivity towards omega-conotoxin GVIA, the non-L-type channel cannot necessarily be considered as an N-type channel because of its distinct voltage-dependent gating properties. 3. Using 65 mM Ca(2+) as the charge carrier, the L-type channels were recruited at about -40 mV and showed a single-channel conductance of 13 pS. Under similar recording conditions, the non-L-type channels were activated at approximately -60 mV and had a single-channel conductance of approximately 16 pS. 4. The non-L-type channel exhibited at least two fast open time constants (tau(o) = 0.2 and 5 ms). In contrast, the L-type channels showed long openings (tau(o) = approximately 23 ms) that were enhanced by Bay K 8644, in addition to the brief openings (tau(o) = 0.3 and 10 ms). 5. The number of functional channels observed in patches of similar sizes suggests that Ca(2+) channels are expressed singly, in low-density clusters (2-15 channels) and in high-density clusters (20-80 channels). Co-localization of the two channel subtypes was observed in patches containing low-density clusters, but was rare in patches containing high-density clusters. 6. Finally, we confirmed the existence of two distinct Ca(2+) channel subtypes by using immunoblot and immunohistochemical techniques.

Strychnine-insensitive [3H] glycine binding to synaptosomal membranes from the chick retina.

The pharmacology and kinetics of strychnine-insensitive [3H] glycine binding to synaptic membranes from the outer (P1) and the inner (P2) plexiform layers of chick retina was studied. Inhibition curves for glycine, D-serine, 1-amincyclopropanecarboxylic acid (ACPC) and strychnine were analyzed by non-linear regression. Hill slopes for glycine and D-serine were not different from unity, whereas those for ACPC were < 1 in both fractions, revealing heterogeneity of binding sites in these membranes. Non-linear regression analysis of time course and saturation experiments strengthen the idea that [3H] glycine binds to more than one class of sites, with similar affinities at equilibrium. Antagonists of strychnine-insensitive glycine receptors in the CNS did not inhibit [3H] glycine binding to these membranes, which demonstrates that NMDA receptors in the retina have different structural requirements for ligand interaction at these sites. pH affected the specific binding, in agreement with the participation of specific amino acid residues at glycine binding sites on NMDA receptors, and also with functional studies in which the modulation of affinity at this site by protons has been observed. These results support previous studies regarding CPP and MK-801 binding, and provide evidence which indicates that the pharmacophore for glycine and other NMDA-related ligands is distinct for the retina, compared to the CNS, mainly regarding the effects of glycine-site antagonists.