A role for inflammatory mediators in the IL-18 mediated attenuation of LTP in the rat dentate gyrus.

Pro-inflammatory cytokines are known to be elevated in several pathological conditions that are associated with deficits in cognition. We have previously demonstrated that interleukin-18 (IL-18) inhibits long-term potentiation (LTP) in the dentate gyrus in vitro. In this study we have examined the involvement of the inflammatory mediators COX-2 and iNOS in IL-18-mediated inhibition of LTP. The effect of an anti-inflammatory PPARgamma agonist was also investigated. We report that the impairment of LTP by IL-18 is significantly attenuated by prior application of the COX-2 inhibitor, SC-236 and the iNOS inhibitor 1400W. These agents had no effect on paired pulse depression in the dentate gyrus. Furthermore, application of the PPARgamma agonist ciglitazone also attenuated IL-18-mediated inhibition of LTP. We discuss a role for p38 MAP kinase in these effects. This study provides novel evidence for the involvement of inflammatory mediators in IL-18-mediated inhibition of LTP in the rat dentate gyrus in vitro.

A role for c-Jun N-terminal kinase in the inhibition of long-term potentiation by interleukin-1beta and long-term depression in the rat dentate gyrus in vitro.

Recent evidence has emphasised the importance of mitogen-activated protein kinase activation in the modulation of hippocampal synaptic plasticity. Whilst extracellular-regulated kinase activation is now regarded as a critical step in the induction of long-term potentiation (LTP), activation of p38 and c-Jun N-terminal kinase (JNK) is associated with its inhibition. Here, the effects of the novel JNK inhibitor anthra[1,9-cd]pyrazol-6(2H)-1 (SP600125) were investigated on the inhibition of LTP by cytokines interleukin-1beta, interleukin-18 and tumour necrosis factor-alpha in the dentate gyrus. Perfusion of SP600125 alone prior to tetanic stimulation of the medial perforant path did not significantly affect baseline synaptic transmission, post-tetanic potentiation or the magnitude of induced LTP. When SP600125 was perfused onto slices prior to application of cytokines, this resulted in a complete reversal of the cytokine-mediated inhibition of LTP. Moreover, the magnitude of LTP attained in these slices was significantly greater than that obtained in vehicle control slices. Next, we investigated the effects of the JNK inhibitor on the impairment of pharmacologically isolated N-methyl-D-aspartate receptor-mediated potentials (NMDA-EPSPs) by interleukin-18. Whilst not affecting baseline amplitude when perfused alone, prior perfusion of SP600125 alleviated the depressive effect of interleukin-18 on NMDA-EPSPs. Finally, we examined the possibility of JNK involvement in the induction of long-term depression (LTD) in the dentate gyrus. Perfusion of SP600125 prior to low-frequency stimulation of the perforant path resulted in a significant attenuation of induced LTD, which suggests that JNK activation is a critical mediator of LTD in the dentate gyrus. These results directly implicate, for the first time, differential activation of JNK in the modulation of distinct forms of hippocampal synaptic plasticity. Whereas acute over-activation of JNK by pathophysiological concentrations of cytokines is detrimental to LTP, physiologic activation of JNK appears necessary for the induction of LTD.

Subtle alterations in growth medium composition can dramatically alter the percentage of unsaturated fatty acids in the yeast Saccharomyces cerevisiae.

The essence of the scientific method is the production of reproducible results in repeated experiments. Cells of the yeast strain DBY747 normally contain 36% unsaturated fatty acids but suddenly, and initially inexplicably, lipid analysis revealed 72% unsaturated fatty acids in the same strain at the same growth temperature. A comparative lipid analysis of DBY747 grown in YEPD and in a number of different types and batches of Yeast Nitrogen Base media revealed two heretofore unreported phenomena. We provide mass spectroscopy and yeast bioassay evidence suggesting that the increase in lipid unsaturation can be attributed to the presence of the plasticizing agent dioctylphthalate in YNB and bactopeptone packaged in 'new' plastic containers first introduced by Difco some 3-4 years ago. We also demonstrate that L-methionine plays an important role in determining the percentage of unsaturated fatty acids in cells grown in laboratory-produced YNB. The results illustrate a novel aspect of methionine metabolism while at the same time highlighting the need for more stringent control to be exercised by the companies that formulate and package defined media.

Alcohols lower the threshold temperature for the maximal activation of a heat shock expression vector in the yeast Saccharomyces cerevisiae.

When the yeast Saccharomyces cerevisiae is exposed to elevated growth temperatures, genes containing a heat shock element (HSE) in their promoters are activated. This study demonstrates that alcohols lower the temperature required for the maximal activation of such a promoter and that the concentration of alcohol required decreases as its hydrophobicity increases. A similar correlation has been found between the members of this alcohol series and their effect on a range of membrane functions. Our results therefore indicate that perturbation of the cell membrane may play a role in the heat induced activation of this HSE-containing promoter.

When a glycolytic gene on a yeast 2 mu ORI-STB plasmid is made essential for growth its expression level is a major determinant of plasmid copy number.

This study demonstrates how varying the promoter strength of an essential gene on a yeast 2 mu ORI-STB YEp multicopy vector can influence vector copy levels. A phosphoglycerate kinase gene (PGK) on this plasmid was made essential for fermentative growth by transformation into a pgk- yeast strain. When in these PGK+ transformants the requirement for PGK expression was the sole selective criterion for plasmid maintenance, PGK promoter activity was inversely related to vector copy levels. Plasmids with an efficiently-transcribed PGK gene were maintained at approximately one copy per cell, whereas those lacking the UAS that normally directs high basal PGK transcription levels were present at up to 10-15 copies. All cultures of these PGK+ transformants contained only a low proportion of pgk- cells. Since mitotic loss of the plasmid arrests growth through loss of a functional PGK allele, PGK confers high stability to the YEp vector in such a pgk- genetic background. In this system YEp vector levels are probably influenced by PGK transcription because high expression of PGK is needed in rapid fermentative growth. Remarkably, low plasmid PGK promoter activity caused PGK mRNA levels slightly higher than those found in yeast with normal PGK regulation. A higher plasmid copy number is therefore not the only factor counteracting the effects of low PGK transcription, and it is possible that PGK mRNA becomes more stable in response to inefficient PGK transcription.

Alpha-factor enhancement of hybrid formation by protoplast fusion in Saccharomyces cerevisiae II.

When Mat a cells are treated with alpha-factor prior to being protoplasted and fused, the frequency of karyogamy is higher than in unarrested controls.

Alpha-factor enhancement of hybrid formation by protoplast fusion in the yeast Saccharomyces cerevisiae.

Two a strains of Saccharomyces cerevisiae, carrying complementary genetic markers, were arrested for 3 h with alpha-factor. These were then protoplasted, prior to fusion using polyethylene glycol. The number of viable fusion products was enhanced by a factor of 20 as compared with unarrested controls.