Potentiation of the glutamatergic synaptic input to rat locus coeruleus neurons by P2X7 receptors

Locus coeruleus (LC) neurons in a rat brain slice preparation were superfused with a Mg²⁺-free and bicuculline-containing external medium. Under these conditions, glutamatergic spontaneous excitatory postsynaptic currents (sEPSCs) were recorded by means of the whole-cell patch-clamp method. ATP, as well as its structural analogue 2-methylthio ATP (2-MeSATP), both caused transient inward currents, which were outlasted by an increase in the frequency but not the amplitude of the sEPSCs. PPADS, but not suramin or reactive blue 2 counteracted both effects of 2-MeSATP. By contrast, α,β-methylene ATP (α,β-meATP), UTP and BzATP did not cause an inward current response. Of these latter agonists, only BzATP slightly facilitated the sEPSC amplitude and strongly potentiated its frequency. PPADS and Brilliant Blue G, as well as fluorocitric acid and aminoadipic acid prevented the activity of BzATP. Furthermore, BzATP caused a similar facilitation of the miniature (m)EPSC (recorded in the presence of tetrodotoxin) and sEPSC frequencies (recorded in its absence). Eventually, capsaicin augmented the frequency of the sEPSCs in a capsazepine-, but not PPADS-antagonizable, manner. In conclusion, the stimulation of astrocytic P2X7 receptors appears to lead to the outflow of a signalling molecule, which presynaptically increases the spontaneous release of glutamate onto LC neurons from their afferent fibre tracts. It is suggested, that the two algogenic compounds ATP and capsaicin utilise separate receptor systems to potentiate the release of glutamate and in consequence to increase the excitability of LC neurons.     

Dual effect of acid pH on purinergic P2X3 receptors depends on the histidine 206 residue

Whole cell patch clamp investigations were carried out to clarify the pH sensitivity of native and recombinant P2X(3) receptors. In HEK293 cells permanently transfected with human (h) P2X(3) receptors (HEK293-hP2X(3) cells), an acidic pH shifted the concentration-response curve for alpha,beta-methylene ATP (alpha,beta-meATP) to the right and increased its maximum. An alkalic pH did not alter the effect of alpha,beta-meATP. Further, a low pH value increased the activation time constant (tau(on)) of the alpha,beta-meATP current; the fast and slow time constants of desensitization (tau(des1), tau(des2)) were at the same time also increased. Finally, acidification accelerated the recovery of P2X(3) receptors from the desensitized state. Replacement of histidine 206, but not histidine 45, by alanine abolished the pH-induced effects on hP2X(3) receptors transiently expressed in HEK293 cells. Changes in the intracellular pH had no effect on the amplitude or time course of the alpha,beta-meATP currents. The voltage sensitivity and reversal potential of the currents activated by alpha,beta-meATP were unaffected by extracellular acidification. Similar effects were observed in a subpopulation of rat dorsal root ganglion neurons expressing homomeric P2X(3) receptor channels. It is suggested that acidification may have a dual effect on P2X(3) channels, by decreasing the current amplitude at low agonist concentrations (because of a decrease in the rate of activation) and increasing it at high concentrations (because of a decrease in the rate of desensitization). Thereby, a differential regulation of pain sensation during e.g. inflammation may occur at the C fiber terminals of small DRG neurons in peripheral tissues.     

Natural selection on ecophysiological traits of a fern species in a temperate rainforest

Unlike other species of the genus Blechnum, the fern Blechnum chilense occurs in a wide range of habitats in Chilean temperate rainforest, from shaded forest understories to abandoned clearings and large gaps. We asked if contrasting light environments can exert differential selection on ecophysiological traits of B. chilense. We measured phenotypic selection on functional traits related to carbon gain: photosynthetic capacity (A _max), dark respiration rate (R _d), water use efficiency (WUE), leaf size and leaf thickness in populations growing in gaps and understorey environments. We assessed survival until reproductive stage and fecundity (sporangia production) as fitness components. In order to determine the potential evolutionary response of traits under selection, we estimated the genetic variation of these traits from clonally propagated individuals in common garden experiments. In gaps, survival of B. chilense was positively correlated with WUE and negatively correlated with leaf size. In contrast, survival in shaded understories was positively correlated with leaf size. We found positive directional fecundity selection on WUE in gaps population. In understories, ferns of lower R _d and greater leaf size showed greater fecundity. Thus, whereas control of water loss was optimized in gaps, light capture and net carbon balance were optimized in shaded understories. We found a significant genetic component of variation in WUE, R _d and leaf size. This study shows the potential for evolutionary responses to heterogeneous light environments in functional traits of B. chilense, a unique fern species able to occupy a broad successional niche in Chilean temperate rainforest.     

Outcrossing increases infection success in the holoparasitic mistletoe <Emphasis Type="Italic">Tristerix aphyllus</Emphasis> (Loranthaceae)

Most studies on the fitness advantage of outbreeding in host–parasite systems have been assessed from the host rather than the parasite perspective. Here, we performed experimental pollination treatments to evaluate the consequences of outbreeding on fitness-related traits in the holoparasitic mistletoe Tristerix aphyllus in a 2-year field study. Results indicate that self-pollinated plants had a lower fruit production than outcrossed plants (20.4% and 29.5% reduction in 2002 and 2003, respectively), and resulting inbred fruits were smaller than outcrossed fruits in both years. No effect was detected for seed mass. The percentage of germination of inbred seeds was 15.1% and 6.0% lower than outcrossed seeds in 2002 and 2003, respectively. Inbred seedlings had shorter radicles, which translated to a 71.6% and 60.0% reduction in infection success compared with outcrossed plants in 2002 and 2003, respectively. Overall, our results revealed significant inbreeding depression on almost every trait that was examined. Although the mean value of traits varied from a year to another, the magnitude of inbreeding depression did not change significantly between years. Our findings constitute the first evidence that outcrossing increases infection success and probably virulence in parasitic plant populations.     

Nocistatin and nociceptin given centrally induce opioid-mediated gastric mucosal protection

Nociceptin (N/OFQ) and nocistatin (NST) are two endogenous neuropeptides derived from the same precursor protein, preproN/OFQ. The aim of the present work was to study the effect of NST on the ethanol-induced mucosal damage compared with that of N/OFQ following intracerebroventricular (i.c.v.) administration in the rat and to analyze the mechanism of the gastroprotective action. It was found that both NST and N/OFQ reduced the mucosal lesions in the same dose range (0.2–1 nmol i.c.v.), but in higher doses (2–5 nmol i.c.v.) the gastroprotective effect of both peptides was highly diminished. The gastroprotective effect of N/OFQ (1 nmol), but not that of NST (1 nmol), was reduced by the selective nociceptin receptor antagonist J-113397 (69 nmol i.c.v.). Similarly, decrease of the gastroprotective effect was observed after the combination of NST (1 nmol) with N/OFQ (0.6 or 1 nmol). However, addition of the gastroprotective effects was observed, when lower dose (0.2 nmol) of NST was given prior to N/OFQ (0.6 nmol). The gastroprotective effect of both N/OFQ and NST was antagonized by naloxone (27 nmol), β-funaltrexamine (20 nmol), naltrindole (5 nmol) and norbinaltorphimine (14 nmol), the μ-, δ- and κ-opioid receptor antagonists, respectively, given i.c.v. The mucosal protection was significantly decreased after bilateral cervical vagotomy. The present findings suggest that NST similar to N/OFQ, may also induce gastric mucosal protective action initiated centrally in a vagal-dependent mechanism. Opioid component is likely to be involved in the gastroprotective effect of both NST and N/OFQ.     

RNA removal from Candida utilis by chemical and thermal treatments

The removal of the content of nucleic acids of fodder yeast (Candida utilis) by treatment with HCl or heat shock was investigated Acid concentrations between 5 and 15% (on dried matter basis) were used. A maximal removal of the content of nucleic acids of 88% was realized wheńn was worked with 15% HCl, 90 °C during 30 minutes. But under this conditions were observed high demanges of protein and also of some essential amino acids Good results for diminishing the content of nucleic acids were reached with the highest concentration of acid and a treatment during 20 minutes The experiments with heat shock were carried out at 68 °C, different times for heating and different contents of yeast In this way better results than for treatment with acid according to diminishing the content of nucleic acids and yield of protein and essential amino acids were reached A removal of over 80% of the content of nucleic acids was achieved in all cases.     

Nucleic acid reduction from yeast activation of intracellular rnase

The study of a heat-shock process for RNA reduction was carried out for different yeast strains. Different results were obtained from each of them. Candida utilis NRRL Y-660 shows its best performance after a 8-s. heat-shock in the presence of 3% NaCl. For commercial baker's yeast Saccharomyces cerevisiae and Kluyveromyces fragilis L-1930, similar results were obtained with only 1% of NaCl. The latter needed longer heat-shock periods. e.g. 15s. to give such an RNA reduction. Biomass recovery ranged from 60 to 75%, being higher for C. utilis and K. fragilis while excessive losses were observed in S. cerevisiae cells. No significant protein deterioration was obtained in the best performance samples. The aminoacid profile appears to be improved in comparison to the starting material in these strains after RNA reduction.     

Host plant and natural enemy impact on cereal aphid competition in a seasonal environment

Competition between phytophagous insects could be mediated by differential impact of natural enemies or by induced changes in host plant quality. The interaction between the aphids Sipha flava and Rhopalosiphum maidis on the shared host plant, Sorghum halepense , was evaluated during the fall and spring seasons in the presence or absence of natural enemies in a garden experiment. During the fall, S. flava was negatively affected by R. maidis in all treatments. However, during the spring season, S. flava was not affected by R. maidis when natural enemies were excluded, and positively affected by R. maidis with natural enemies present. Rhopalosiphum maidis was negatively affected by S. flava during the fall and spring seasons under all treatments. Laboratory experiments demonstrated that changes in host plant quality induced by S. flava attack and the presence of the parasitoid Lysiphlebus testaceipes negatively affected R. maidis , but not S. flava . Under abiotic conditions simulating fall and spring (lower temperature and daylength in the fall than in the spring), S. flava showed lower performance than R. maidis under "fall" conditions, and both aphid species showed similar life history traits under "spring" conditions. For S. flava , the time to the first reproduction and longevity were longer, and the nymphal production was lower, under "fall" conditions than under "spring" conditions. Our results showed that S. flava is benefited when R. maidis and the parasitoid L. testaceipes are present but the abiotic conditions are not adverse.