Interplay mechanisms between progesterone and endocannabinoid receptors in regulating bull sperm capacitation and acrosome reaction.

After ejaculation, sperm show a limited capacity for transcription and translation. In the oviduct, most of the signalling in sperm is nongenomic and is mediated through membrane receptors. Studies have shown that the cation channel of sperm (CatSper), cAMP, cGMP, protein kinases, and tyrosine phosphorylation are involved in the nongenomic signalling of progesterone (P4) in sperm. However, it is not known whether there is an interplay between P4 and cannabinoid receptors 1 and 2 (CB1 and CB2), transient receptor potential vanilloid 1 (TRPV1), CatSper channels, cAMP, inositol trisphosphate receptor (IP3R), and mitogen-activated protein kinase (MAPK); these potential regulators are involved in the regulation of capacitation and the acrosome reaction. In the present study, selective blockers of CB1, CB2, TRPV1, CatSper channels, cAMP, protein kinase A (PKA), IP3R, and MAPK were used to identify their involvement in P4-mediated bull sperm capacitation and the acrosome reaction. Selective blocking of any one of the molecules caused a significant reduction in P4 signalling (p < 0.05). Interestingly, blocking these molecules in combination followed by treatment with P4 resulted in the complete absence of capacitation and the acrosome reaction. Blocking a single receptor was not able to eliminate the P4-induced capacitation and the acrosome reaction. In addition to the CB1 and CB2 receptors, there may be other signalling pathways that mediate P4 signalling. In conclusion, P4 signalling exhibited interplay with the cannabinoid receptors. The regulation of sperm capacitation and the acrosome reaction also involved cAMP, PKA, l-type and T-type calcium channels, TRPV1, inositol trisphosphate, and MAPK.

Effect of micronutrient supplementation around calving on the plasma cortisol levels of Murrah buffaloes and Sahiwal and Karan Fries cows.

Micronutrients when fed around peripartum may reduce the stress induced by cortisol. With this objective, 24 Sahiwal (SW) and 24 Karan Fries (KF) cows and 24 Murrah (Mu) buffaloes were taken and divided into four groups of six each. Vitamin E (VE), zinc (Zn) and copper were supplemented from 30 days pre- to 30 days postcalving in groups 1, 2 and 3. Animals without supplementation served as control. Blood sampling was done on days 30, 15, 7 and 3 precalving; at calving; and on days 3, 7, 15 and 30 postcalving. Plasma cortisol levels were measured by ELISA. Supplementation of VE and Zn significantly (P<0.05) reduced plasma cortisol levels at calving and at 30 days postcalving. KF cows exhibited best results with VE, whereas SW cows and Mu buffaloes responded best to Zn. Copper increased the peripartum cortisol levels. Reduction in cortisol levels may help in keeping the animal healthy and stronger to fight immuno-suppression generally observed around the period of peripartum.

Ameliorative potential of α-tocopherol against flubendiamide and copper-induced testicular-insult in Wistar rats.

Study was undertaken to evaluate ameliorative potential of α-tocopherol against copper sulphate and flubendiamide alone and in combination-induced toxicity in rats following 90 days exposure. Absolute and relative organ weights did not differ between treatments groups. Increase of LPO in copper and flubendiamide intoxicated rats but modest increase in copper + flubendiamide group. GSH and activities of SOD, GPx and GST showed moderate decrease in intoxicated groups. Reduced CAT activity in alone exposed groups was observed. ACP, ALP and SDH remain unaltered. Increase in LDH, γ-GT, abnormal sperm and reduced 17ß-HSD, percent live and HOST +ve sperms and testosterone level was observed in all three exposed groups. Xenobiotics alone and in combination exhibited degenerative germinal epithelium, necrotic germ cells, loss of spermatozoa and spermatids. Treatment with α-tocopherol, reparative potential was observed as values of most of the parameters including testicular histoarchitecture were restored.

Neutrophil dynamics in the blood and milk of crossbred cows naturally infected with Staphylococcus aureus.

AIM: The present study was designed to evaluate the neutrophil dynamics in terms of the functional competence during subclinical mastitis (SCM) and clinical mastitis (CM). MATERIALS AND METHODS: A total of 146 Karan fries cows were screened and were divided into three groups as control (n=12), SCM, n=12 and CM, n=12 groups on the basis of California mastitis test scoring, bacteriological evaluation, gross and morphological changes in milk and by counting milk somatic cell count (SCC). Both blood and milk polymorphonuclear neutrophils (PMNs) were isolated in the study. Phagocytic activity (PA) was studied by spectrophotometrically; neutrophil extracelluar traps (NETs) were studied by scanning electron microscopy (SEM); CD44 was quantified by flow cytometry and apoptosis was studied by fluorescent microscopy. RESULTS: Significantly (p<0.05) higher SCC, PA was found in milk of CM cows as compared to SCM and control cows. Significantly lower (p<0.05) apoptosis was observed in PMNs isolated from both blood and milk of CM group of cows when compared to control and SCM group. The milk neutrophils of CM group of cows formed NETs as evidenced from the SEM images. Surface expression of CD44 revealed a significantly (p<0.05) lower expression in milk neutrophils of CM group of cows when compared to SCM and control group of cows. CONCLUSION: The study indicated a positive correlation between delayed neutrophil apoptosis, persistent staying of neutrophils at the site of infection along with formation of NETs as the strategies to fight against the pathogens in the udder during Staphylococcal mastitis. The study forms a strong base for future molecular research in terms of neutrophil recruitment and neutrophil removal from the site of infection.

Formation of NET, phagocytic activity, surface architecture, apoptosis and expression of toll like receptors 2 and 4 (TLR2 and TLR4) in neutrophils of mastitic cows.

Neutrophils employ both oxidative and non oxidative mechanisms to destroy pathogens. Function of neutrophils coming in milk during mammary invasion is not clearly understood in dairy animals. Therefore, the present study was designed in 36 Sahiwal cows to see the changes occurring in the neutrophil activity of cows suffering from subclinical (SCM) and clinical mastitis (CM). Cows were divided into three groups as healthy (n = 12), SCM (n = 12) and CM (n = 12) groups on the basis of CMT scoring, gross morphological changes in milk, bacteriological examination of milk and by counting milk SCC. Significantly (P < 0.05) higher milk SCC, neutrophil percent and significantly (P < 0.05) lower viability of both blood and milk neutrophils were observed in CM group of cows as compared to SCM and control group of cows. Phagocytic activity (PA) was significantly (P < 0.05) higher in milk neutrophils of SCM and CM cows as compared to control cows. Toll like receptors 2 and 4 in blood and milk neutrophils were found to be significantly (P < 0.05) higher, whereas, apoptosis in terms of altered mitochondrial transmembrane potential, Caspase 3 and 7 activities were found to be significantly (P < 0.05) lower in CM cows. Alterations in surface architecture of neutrophils in terms of formation of pseudopods was observed by scanning electron microscope (SEM) and found to be higher in CM cows. Blood neutrophils were found to be spherical as compared to milk neutrophils. Formation of neutrophil extracellular trap (NETs) were found milk neutrophils of CM cows, whereas, SCM and healthy cows did not exhibit NET formation. The study indicated a positive correlation between lower neutrophil apoptosis and higher expression of TLR2 and TLR4 with the formation of NETs and change in surface architecture. Formation of NET like structures seemed to be an effective mode of defense employed by neutrophils of cows suffering from clinical mastitis.