The "Ram Effect": A "Non-Classical" Mechanism for Inducing LH Surges in Sheep

During spring sheep do not normally ovulate but exposure to a ram can induce ovulation. In some ewes an LH surge is induced immediately after exposure to a ram thus raising questions about the control of this precocious LH surge. Our first aim was to determine the plasma concentrations of oestradiol (E2) E2 in anoestrous ewes before and after the "ram effect" in ewes that had a "precocious" LH surge (starting within 6 hours), a "normal" surge (between 6 and 28h) and "late» surge (not detected by 56h). In another experiment we tested if a small increase in circulating E2 could induce an LH surge in anoestrus ewes. The concentration of E2 significantly was not different at the time of ram introduction among ewes with the three types of LH surge. "Precocious" LH surges were not preceded by a large increase in E2 unlike "normal" surges and small elevations of circulating E2 alone were unable to induce LH surges. These results show that the "precocious" LH surge was not the result of E2 positive feedback. Our second aim was to test if noradrenaline (NA) is involved in the LH response to the "ram effect". Using double labelling for Fos and tyrosine hydroxylase (TH) we showed that exposure of anoestrous ewes to a ram induced a higher density of cells positive for both in the A1 nucleus and the Locus Coeruleus complex compared to unstimulated controls. Finally, the administration by retrodialysis into the preoptic area, of NA increased the proportion of ewes with an LH response to ram odor whereas treatment with the α1 antagonist Prazosin decreased the LH pulse frequency and amplitude induced by a sexually active ram. Collectively these results suggest that in anoestrous ewes NA is involved in ram-induced LH secretion as observed in other induced ovulators

The effect of an intracerebroventricular injection of metformin or AICAR on the plasma concentrations of melatonin in the ewe: potential involvement of AMPK?

<p>Abstract</p> <p>Background</p> <p>It is now widely accepted that AMP-activated protein kinase (AMPK) is a critical regulator of energy homeostasis. Recently, it has been shown to regulate circadian clocks. In seasonal breeding species such as sheep, the circadian clock controls the secretion of an endogenous rhythm of melatonin and, as a consequence, is probably involved in the generation of seasonal rhythms of reproduction. Considering this, we identified the presence of the subunits of AMPK in different hypothalamic nuclei involved in the pre- and post-pineal pathways that control seasonality of reproduction in the ewe and we investigated if the intracerebroventricular (i.c.v.) injection of two activators of AMPK, metformin and AICAR, affected the circadian rhythm of melatonin in ewes that were housed in constant darkness. In parallel the secretion of insulin was monitored as a peripheral metabolic marker. We also investigated the effects of i.c.v. AICAR on the phosphorylation of AMPK and acetyl-CoA carboxylase (ACC), a downstream target of AMPK, in brain structures along the photoneuroendocrine pathway to the pineal gland.</p> <p>Results</p> <p>All the subunits of AMPK that we studied were identified in all brain areas that were dissected but with some differences in their level of expression among structures. Metformin and AICAR both reduced (p < 0.001 and p < 0.01 respectively) the amplitude of the circadian rhythm of melatonin secretion independently of insulin secretion. The i.c.v. injection of AICAR only tended (p = 0.1) to increase the levels of phosphorylated AMPK in the paraventricular nucleus but significantly increased the levels of phosphorylated ACC in the paraventricular nucleus (p < 0.001) and in the pineal gland (p < 0.05).</p> <p>Conclusions</p> <p>Taken together, these results suggest a potential role for AMPK on the secretion of melatonin probably acting trough the paraventricular nucleus and/or directly in the pineal gland. We conclude that AMPK may act as a metabolic cue to modulate the rhythm of melatonin secretion.</p

Five omic technologies are concordant in differentiating the biochemical characteristics of the berries of five grapevine (Vitis vinifera L.) cultivars

Background: Grape cultivars and wines are distinguishable by their color, flavor and aroma profiles. Omic analyses (transcripts, proteins and metabolites) are powerful tools for assessing biochemical differences in biological systems. Results: Berry skins of red- (Cabernet Sauvignon, Merlot, Pinot Noir) and white-skinned (Chardonnay, Semillon) wine grapes were harvested near optimum maturity (°Brix-to-titratable acidity ratio) from the same experimental vineyard. The cultivars were exposed to a mild, seasonal water-deficit treatment from fruit set until harvest in 2011. Identical sample aliquots were analyzed for transcripts by grapevine whole-genome oligonucleotide microarray and RNAseq technologies, proteins by nano-liquid chromatography-mass spectroscopy, and metabolites by gas chromatography-mass spectroscopy and liquid chromatography-mass spectroscopy. Principal components analysis of each of five Omic technologies showed similar results across cultivars in all Omic datasets. Comparison of the processed data of genes mapped in RNAseq and microarray data revealed a strong Pearson's correlation (0.80). The exclusion of probesets associated with genes with potential for cross-hybridization on the microarray improved the correlation to 0.93. The overall concordance of protein with transcript data was low with a Pearson's correlation of 0.27 and 0.24 for the RNAseq and microarray data, respectively. Integration of metabolite with protein and transcript data produced an expected model of phenylpropanoid biosynthesis, which distinguished red from white grapes, yet provided detail of individual cultivar differences. The mild water deficit treatment did not significantly alter the abundance of proteins or metabolites measured in the five cultivars, but did have a small effect on gene expression. Conclusions: The five Omic technologies were consistent in distinguishing cultivar variation. There was high concordance between transcriptomic technologies, but generally protein abundance did not correlate well with transcript abundance. The integration of multiple high-throughput Omic datasets revealed complex biochemical variation amongst five cultivars of an ancient and economically important crop species.26 page(s

Additional file 3: of Five omic technologies are concordant in differentiating the biochemical characteristics of the berries of five grapevine (Vitis vinifera L.) cultivars

Annotation, protein spectral counts, Normalized Spectral Abundance Factor (NSAF) values and log2 transformed NSAF values for each replicate and protein identified, with ‘.count’, ‘.NSAF’, and ‘.NSAF.log2’ suffixes, respectively. Mass weight (Mr) and amino acid (AA) length are approximated values. Cultivar and treatment abbreviations for experimental replicates: Cabernet Sauvignon (CS), Merlot (ME), Pinot Noir (PN), Chardonnay (CD), and Semillon (SM) grown under well-watered (W) and water deficit (D) conditions, n = 3. Other column headings refer to a combination of cultivar name, treatment, and replicate number. For example, CDD2, is Chardonnay, water deficit and replicate 2. (XLS 4096 kb

Additional file 9: of Five omic technologies are concordant in differentiating the biochemical characteristics of the berries of five grapevine (Vitis vinifera L.) cultivars

Correlations of protein and transcript abundance. Protein data are log2 NSAF values, n = 3, RNAseq data are log2 normalized counts per million (CPM), n = 3, and microarray data are log2 RMA values, n = 5. Relationships of proteins with either RNAseq (CPM) or microarray (RMA) are indicated. (XLS 370 kb

Additional file 7: of Five omic technologies are concordant in differentiating the biochemical characteristics of the berries of five grapevine (Vitis vinifera L.) cultivars

Mean relative abundance values, M/Z, and results from the ANOVA for all primary and secondary metabolomic details for all metabolites (67) analyzed by GC-MS and (42) analyzed by LC-MS in five grape cultivars. Cultivar and treatment abbreviations for experimental replicates: Cabernet Sauvignon (CS), Merlot (ME), Pinot Noir (PN), Chardonnay (CD), and Semillon (SM) grown under well-watered (W) and water deficit (D) conditions, n = 6. (XLS 76 kb