Interactions between Nutrition and the "Ram Effect" in the Control of Ovarian Function in the Merino Ewe.

We tested whether short-term nutritional supplementation (500 g lupin grain per head daily) would affect the response of ewes to the ram effect. Experiment 1 (end of non-breeding season): ewes were supplemented for either Days -6 to -1 relative to ram introduction (n = 24) or for 12 days after ram introduction (Days 11 to 22 of the ram-induced cycle; n = 29). Controls (n = 30) were not supplemented. Across all groups, 94-100% of ewes ovulated. Supplementation before ram introduction did not affect ovulation rate at the ram-induced ovulation but increased it during the ram-induced cycle (Control 1.37; supplemented 1.66; p < 0.05). Experiment 2 (the middle of non-breeding season): the supplement was fed for Days -5 to -1 relative to ram introduction. Again, supplementation did not increase number ovulating (Control 16/29; Supplemented 10/29) but it did increase ovulation rate at the ram-induced ovulation (Control 1.31; Supplemented 1.68; p < 0.05). In neither experiment did supplementation affect the frequency of short cycles. Supplementation before ram introduction did not improve the percentage of ewes ovulating or reduce the frequency of short cycles (so will not improve the synchrony of lambing). However, supplementation after ram introduction can increase prolificacy.

Roles of small RNAs in the effects of nutrition on apoptosis and spermatogenesis in the adult testis.

We tested whether reductions in spermatozoal quality induced by under-nutrition are associated with increased germ cell apoptosis and disrupted spermatogenesis, and whether these effects are mediated by small RNAs. Groups of 8 male sheep were fed for a 10% increase or 10% decrease in body mass over 65 days. Underfeeding increased the number of apoptotic germ cells (P < 0.05) and increased the expression of apoptosis-related genes (P < 0.05) in testicular tissue. We identified 44 miRNAs and 35 putative piRNAs that were differentially expressed in well-fed and underfed males (FDR < 0.05). Some were related to reproductive system development, apoptosis (miRNAs), and sperm production and quality (piRNAs). Novel-miR-144 (miR-98), was found to target three apoptotic genes (TP53, CASP3, FASL). The proportion of miRNAs as a total of small RNAs was greater in well-fed males than in underfed males (P < 0.05) and was correlated (r = 0.8, P < 0.05) with the proportion of piRNAs in well-fed and underfed males. In conclusion, the reductions in spermatozoal quality induced by under-nutrition are caused, at least partly, by disruptions to Sertoli cell function and increased germ cell apoptosis, mediated by changes in the expression of miRNAs and piRNAs.

Under-nutrition reduces spermatogenic efficiency and sperm velocity, and increases sperm DNA damage in sexually mature male sheep.

We tested whether the quality of spermatozoa from mature male sheep would be affected during nutrition-induced changes in testicular mass. Merino rams were fed for 65 days with diets that increased, maintained or decreased body and testis mass (n=8 per group). In semen collected on Days 56 and 63, underfed rams had less sperms per ejaculate than well-fed rams (P<0.05) and a lower sperm velocity (computer-assisted semen analysis) than well-fed or maintenance-fed rams (P<0.05). Sperm chromatin structure assay revealed more sperm DNA damage in underfed rams than in well-fed rams (P<0.05). The amount of sperm DNA damage was inversely correlated with change in scrotal circumference (r=-0.6, P<0.05), the percentages of progressive motile sperm (r=-0.8; P<0.01) and motile sperm (r=-0.6, P<0.05), and the numbers of sperms per gram of testis (r=-0.55, P<0.05). In testicular tissue collected on Day 65, underfed rams had fewer sperm per gram of testis than rams in the other two groups (P<0.001). We conclude that, in adult rams, underfeeding reduces spermatogenic efficiency and that this response is associated with a reduction in spermatozoal quality.

The 'male effect' in sheep and goats--revisiting the dogmas.

Male-induced ovulation in sheep and goats (the 'male effect'), documented during the period 1940-1960, has long been shrouded in preconceptions concerning how, when and why it worked. These preconceptions became dogmas but recent research is challenging them so, in this review, we have re-visited some major physiological (breed seasonality; characteristics of the response; the nature of the male stimuli) and physical factors (duration of male presence; isolation from male stimuli) that affect the phenomenon. We reject the dogma that ewes must be isolated from males and conclude that male 'novelty' is more important than isolation per se. Similarly, we reject the perception that the neuroendocrine component of the male effect is restricted to anovulatory females. Finally, we re-assess the relative importance of olfactory and non-olfactory signals, and develop a perspective on the way male-induced ovulation fits with preconceptions about pheromonal processes in mammals. Overall, our understanding of the male effect has evolved significantly and it is time to modify or reject our dogmas so this field of research can advance. We can now ask new questions regarding the application of the male effect in industry and develop research so we can fully understand this biological phenomenon.

Rapid induction of cell proliferation in the adult female ungulate brain (Ovis aries) associated with activation of the reproductive axis by exposure to unfamiliar males.

In many species, the reproductive centers of the brain are profoundly affected by sociosexual stimuli. This is particularly evident in female ungulates such as sheep, in which exposure to males switches them from reproductively quiescent to fertile. In two experiments with female sheep, we tested whether the brain centers that control gonadotropin-releasing hormone (GnRH) neuronal activity respond differentially to "novel" vs. familiar males and whether the neuroendocrine response is associated with increased cell proliferation in the hippocampus, a site associated with memory formation. In experiment 1, groups of 10 female sheep that had previously been habituated to males for 3 mo were re-exposed to familiar males or were exposed to novel males. Only the novel males increased luteinizing hormone (LH) pulse frequency, indicating stimulation of GnRH neuronal activity. In experiment 2, groups of six female sheep were injected with bromodeoxyuridine (BrdU) and then maintained in isolation from males or exposed to novel males. Two days later, the hippocampus and hypothalamus were removed and processed for fluorescence immunohistochemistry. Again, exposure to males increased LH pulse frequency. Most important, male exposure also doubled the number of BrdU-positive cells in the dentate gyrus of the hippocampus. No BrdU-positive cells were detected in the hypothalamus. We conclude that the stimulus from novel males switches on the reproductive centers of the brain of female sheep and rapidly doubles the rate of cell proliferation in the hippocampus. The rapidity of this response contrasts with rodents, in which several days of exposure to male pheromones seem necessary for an effect on neurogenesis.