Lead and calcium crosstalk tempted acrosome damage and hyperpolarization of spermatozoa: signaling and ultra-structural evidences.

BACKGROUND: Exposure of humans and animals to heavy metals is increasing day-by-day; thus, lead even today remains of significant public health concern. According to CDC, blood lead reference value (BLRV) ranges from 3.5 µg/dl to 5 µg/dl in adults. Recently, almost 2.6% decline in male fertility per year has been reported but the cause is not well established. Lead (Pb2+) affects the size of testis, semen quality, and secretory functions of prostate. But the molecular mechanism(s) of lead toxicity in sperm cells is not clear. Thus, present study was undertaken to evaluate the adverse effects of lead acetate at environmentally relevant exposure levels (0.5, 5, 10 and 20 ppm) on functional and molecular dynamics of spermatozoa of bucks following in vitro exposure for 15 min and 3 h. RESULTS: Lead significantly decreased motility, viable count, and motion kinematic patterns of spermatozoa like curvilinear velocity, straight-line velocity, average path velocity, beat cross frequency and maximum amplitude of head lateral displacement even at 5 ppm concentration. Pb2+ modulated intracellular cAMP and Ca2+ levels in sperm cells through L-type calcium channels and induced spontaneous or premature acrosome reaction (AR) by increasing tyrosine phosphorylation of sperm proteins and downregulated mitochondrial transmembrane potential. Lead significantly increased DNA damage and apoptosis as well. Electron microscopy studies revealed Pb2+ -induced deleterious effects on plasma membrane of head and acrosome including collapsed cristae in mitochondria. CONCLUSIONS: Pb2+ not only mimics Ca2+ but also affects cellular targets involved in generation of cAMP, mitochondrial transmembrane potential, and ionic exchange. Lead seems to interact with Ca2+ channels because of charge similarity and probably enters the sperm cell through these channels and results in hyperpolarization. Our findings also indicate lead-induced TP and intracellular Ca2+ release in spermatozoa which in turn may be responsible for premature acrosome exocytosis which is essential feature of capacitation for fertilization. Thus, lead seems to reduce the fertilizing capacity of spermatozoa even at 0.5 ppm concentrations.

Quercetin in semen extenders curtails reactive oxygen and nitrogen species and improves functional attributes of cryopreserved buck semen.

Cryopreservation of goat spermatozoa is challenging due to several factors, including one of the most essential, i.e., oxidative stress. It is particularly essential in goat semen due to its scanty ejaculate volume and high sperm concentration. This leaves a narrow sperm-to-seminal plasma ratio owing to marginal antioxidant support; moreover, semen extension further dilutes the antioxidant level, leading to an imbalance of oxidant-antioxidant equilibrium. The present study aimed to evaluate the effect of quercetin on curtailing oxidative stress and its reflection on the post-thaw survivability and membrane integrity of goat spermatozoa. For this study, six bucks were selected. Six ejaculates from each buck totaling 36 ejaculates were collected, which were then split into five parts; furthermore, each part was added with a semen extender having a particular concentration of additive. Group C without quercetin and T1 containing Vit E at 3 mmol/mL were considered the control and positive control respectively, whereas T2, T3, and T4 contain 10, 20, and 30 µmol/mL of Quercetin respectively. The final sperm concentration of each group was kept at 200×106 spermatozoa/mL. All groups were subjected to equilibration at 4 °C for 4 hours, then filled in French mini (0.25 mL) straws, followed by sealing and cryopreservation. Samples after 72 hours of cryopreservation were subjected to evaluation of plasma membrane integrity and viability through staining, acrosomal integrity, and mitochondrial membrane activity through flowcytometry. Evaluation of sperm kinematics as well as the oxidant-antioxidant status of sperm (ROS and nitric oxide) and seminal plasma (SOD, CAT, GPx, FRAP, and lipid peroxidation through MDA estimation) were also carried out. Quercetin, when supplemented at 20 µmol/mL in buck semen extender, significantly (p<0.01) improved cryopreserved sperm functions in terms of plasma membrane integrity, viability, acrosomal integrity, mitochondrial membrane activity, and sperm kinematics of buck semen. Similarly, Quercetin supplementation at 20 µmol/mL significantly reduced reactive oxygen and nitrogen species (RONS) in sperm and improved the antioxidant status of seminal plasma, which was indicated by reduced oxidative damage and improved the antioxidant status of buck semen. In conclusion, Quercetin at 20 µmol/mL reduced oxidative stress, improved semen antioxidant status, and improved sperm membrane integrity and kinematics.

Feeding Astragalus membranaceus Root Improves the Rumen Fermentation Rate in Housed Goats through the Alteration of the Rumen Community Composition.

Although Astragalus membranaceus root (AMR) has been noted as an ingredient in ruminant feed, the impacts of AMR feeding on rumen fermentation and the microbial community structure within the rumen are yet to be evaluated. This study investigated the effects of AMR supplementation on rumen fermentation characteristics and microbial community structures in goats. In two sets of feeding experiments, four Japanese native goats were fed AMR (10 g/kg DM/day/head) for three weeks per experiment. The rumen fluid samples were analyzed using high-performance liquid chromatography for fermentation products and next-generation sequencing for microbial analysis. The rumen fluid samples in the second experiment were also subject to an in vitro anaerobic fermentation test. The results indicated a significant modification, with a higher volatile fatty acid (VFA) content in the rumen fluid of goats in the feeding period than before feeding (p < 0.01). The microbial analysis revealed a significant increase in community diversity (p < 0.05) following AMR feeding, and the rumen bacterial community increased in two families belonging to the order Oscillospirales in Firmicutes (p < 0.05). The phylum Verrucomicrobiota was observed to be significantly less abundant after AMR feeding than during the control period (p < 0.05). Notably, the linear discriminant analysis revealed that the families with largely unknown functions in the rumen (Oscillospiraceae, Rikenellaceae, Muribaculaceae, and vadinBB97) were the determinants of the community split between control and AMR feeding. Increased fermentation rate by AMR feeding was also supported by an in vitro culture experiment, which resulted in faster VFA production without affecting methane production in total gas production. The study demonstrated that AMR can significantly facilitate change in the bacterial community structure in the goat rumen involving a shift of the favoring fibrolytic bacteria towards VFA production. The long-term effects of AMR supplementation and its applicability across different ruminant species, with potential benefits for animal health and productivity, should be addressed.

Effect of Current Waveforms during Directed Energy Deposition of 4043 Aluminum Alloy on Microstructure, Hardness, and Wear of Alloy.

Wire arc additive manufacturing (WAAM) was employed to fabricate 4043 aluminum alloy walls. To investigate the effects of sinusoidal, triangular, and rectangular waveforms of alternating current (AC) and their transients on the wall geometry, microstructure evolution, hardness, and wear properties were evaluated. The root mean square (RMS) current value was maximum for the rectangular and minimum for the triangular waveform. The section produced by the triangular waveform had the highest height-to-width ratio, indicating that this waveform can be a favorable choice for creating components using WAAM. The optical micrographs of the transverse cross-section of the printed sections revealed the grain structure produced with this waveform to be heterogeneous, having a columnar dendritic structure at the bottom and equiaxed at the top portion. The waveforms also had an impact on the hardness and wear characteristics of all the walls, which were attributed to their cooling rate.

IL1ß/ TNFα/COX-2/VEGF axis responsible for effective healing potential of C-glucoside xanthone (mangiferin) based ointment in immunocompromised rats.

Present study was conducted to undermine the wound healing potential of mangiferin vis a vis its molecular dynamics in immunocompromised excisional rat model. 120 rats were randomly and equally divided into five groups viz. group I (Healthy control), group II (Immunocompromised control), group III (Immunocompromised group treated with silver sulphadiazine), group IV (Immunocompromised group treated with 2.5 %Mangiferin) and group V (Immunocompromised group treated with 5 %Mangiferin). Immuno compromised state was achieved following intramuscular injection of Hydrocortisone @ 80 mg/kg body weight. Study was conducted for a period of 28 days. Six animals from each group were humanely sacrificed at weekly interval till day 28th of study. Planimetric analysis, biochemical studies viz. hydroxyproline assay, total protein and DNA content, antioxidative potential through LPO assay was done along with molecular studies involving expression profiling of IL1ß, TNFα and COX-2 and Immunohistochemistry of angiogenic marker i.e. VEGF was performed to undermine the pharmacodynamics of mangiferin. Histopathological studies including H&E and Masson's Trichome was also performed to study histoarchitectural changes in wound healing and reparative process following application of mangiferin ointment. Study revealed significant (P ≤ 0.05) reduction in wound area measurement and significant (P ≤ 0.05) increase in wound contraction (%) following mangiferin administration in immunocompromised rats. Hydroxyproline, DNA and total protein showed significant (P ≤ 0.05) increase in skin tissues of mangiferin treated immunocompromised rats. LPO assay revealed significant (P ≤ 0.05) reduction in mangiferin treated animals. Histopathological studies of skin tissues revealed complete restoration advocating grade III of healing in 2.5% mangiferin treated group. Higher expression and strong signal intensity of VEGF was noticed in 2.5% mangiferin treatment group along with significant (P ≤ 0.05) upregulation IL1ß and TNFα on day 7 in 2.5% mangiferin treatment group with significant (P ≤ 0.05) down regulation of COX-2 in mangiferin treatment group as compared to other groups i.e. group II and III. It is concluded from our study that mangiferin facilitates wound healing through improved wound closure, organized deposition of collagen deposition and granulation matrix formation.

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.

Collapsed mitochondrial cristae in goat spermatozoa due to mercury result in lethality and compromised motility along with altered kinematic patterns.

Earlier we have reported mercury-induced alterations in functional dynamics of buck spermatozoa through free radicals-mediated oxidative stress and spontaneous acrosome reaction. Based on our earlier findings, we aimed to investigate the effect of mercury exposure on motility, kinematic patterns, DNA damage, apoptosis and ultra-structural alterations in goat spermatozoa following in vitro exposure to different concentrations (0.031-1.25 µg/ml) of mercuric chloride for 15 min and 3 h. Following exposure of sperm cells to 0.031 µg/ml of mercuric chloride for 3 h, livability and motility of sperms was significantly reduced along with altered kinematic patterns, significant increase in per cent necrotic sperm cells and number of cells showing DNA damage; and this effect was dose- and time-dependent. Contrary to up-regulation of Bax gene after 3 h in control group, there was significant increase in expression of Bcl-2 in mercury-treated groups. Transmission electron microscopy studies revealed rifts and nicks in plasma and acrosomal membrane, mitochondrial sheath, and collapsed mitochondria with loss of helical organization of mitochondria in the middle piece of spermatozoa. Our findings evidently suggest that mercury induces necrosis instead of apoptosis and targets the membrane, acrosome, mid piece of sperms; and the damage to mitochondria seems to be responsible for alterations in functional and kinematic attributes of spermatozoa.

A review on phytochemistry and pharmacological uses of Tecoma stans (L.) Juss. ex Kunth.

ETHNOPHARMACOLOGICAL RELEVANCE: Tecoma stans (L.) Juss. ex Kunth (Bignoniaceae) is an attractive evergreen plant known as kusi urakame, koyawari, Palo amarillo, tronadora, yellow-elder, yellow trumpet bush, trumpet-flower, yellow-bells, trumpet bush, ginger-Thomas, esperanza, and timboco. It is widely used in traditional Mexican medicine, to treat hyperglycemia, gastrointestinal and urinary tract disorders, jaundice, toothaches, headaches, colds, skin infections, and scorpion, snake, and rat bites. Current research focusses on evaluating its bioactive components and therapeutic potential. AIM OF THE STUDY: The current article reviewed the information available on Tecoma stans ethnopharmacology, geographical distribution, chemical composition, phytochemistry, therapeutic effects, and toxicology. MATERIAL AND METHODS: Information of botanical description, distribution, traditional uses, chemical composition, bioactive components, and therapeutic investigations was gathered from a comprehensive literature search of electronic databases such as Science Direct, PubMed, Web of Science, Wiley, ACS, Springer, Taylor and Francis, Google Scholar, and SCOPUS until 2020 for publications (peer-reviewed articles, eBooks, short communications, reports from international organizations, and case letters). Information was also included from books, conference proceedings, and thesis. Primary keywords for data collection were "Tecoma stans," and "Ethnopharmacology," followed by secondary keywords such as "Constituents," "Therapeutic effect," and "Toxicity." RESULTS: An exhaustive comparative study of the accessible sources of Tecoma stans confirmed its origin, ethnopharmacological and therapeutic uses. More than 120 chemical compounds have been isolated, and the main active principles are alkaloids, phenolic acids, flavonoids, and fatty acids. The plant possesses vast therapeutic benefits, such as lowering elevated blood sugar levels, anti-inflammatory, anti-cancer, anti-bacterial, anti-fungal, anti-oxidant, hepatoprotective, and wound healing actions. CONCLUSIONS: Comprehensive literature analysis exhibits that many populations have utilized Tecoma stans around the globe with specific reference to different parts of Mexico. The above information shows that the plant holds many hidden potentials and can, therefore, be studied extensively for its phytoconstituents and therapeutic effects. However, while going through the literature, it was observed that incomplete data is reported on in vivo trials, especially concerning its dosage, positive and negative control groups, intervention time, and toxicity studies. Additionally, there is a lack of information on its complete nutritional and phytochemical profiling. We trust that this review will help lay the groundwork for encouraging pharmacological and pharmaceutical studies. It will also direct us to understand the clinical relevance and applications of bioactive compounds from Tecoma stans in the prevention and treatment of diseases.

Freezing-thawing induces deprotamination, cryocapacitation-associated changes; DNA fragmentation; and reduced progesterone sensitivity in buck spermatozoa.

In the present study, there was evaluation of cryocapacitation-associated changes, apoptotic-like changes, deprotamination, total antioxidant capacity (TAC), and in vitro sperm functional attributes in Barbari bucks after freezing-thawing. The correlation between deprotamination and sperm functional characteristics was established. Using immunoblotting procedures, there was detection of the presence of a single 28-kDa protein band corresponding to protamine-1. The localization in the head region of the spermatozoa was further validated by an immunofluorescence test. Capacitated (B-) and acrosome-reacted (AR-) pattern spermatozoa, spermatozoa with the externalization of phosphatidylserine and a relatively lesser mitochondrial transmembrane potential, and deprotamination and DNA fragmentation was greater (P < 0.05) after freezing-thawing and indicated there were cryocapacitation- and apoptotic-like changes, respectively. Furthermore, the detection of phosphorylation of tyrosine-containing proteins with use of immunoblotting and immunofluorescence procedures confirmed there were cryocapacitation-like changes in the buck spermatozoa after freezing-thawing. Total antioxidant capacity (TAC), in vitro thermal resistance response, Vanguard distance, progesterone sensitivity, and in vitro capacitation response were less (P < 0.05) in the spermatozoa after freezing-thawing compared with spermatozoa after initial dilution and equilibration. Deprotamination (chromomycin A3-positive cells, CMA3+) and DNA fragmentation (TUNEL+ve) were positively correlated with B- and AR-pattern spermatozoa, while other values for other variables were negatively correlated. In conclusion, the results of this study indicated there was protamine-1 in buck spermatozoa and after freezing-thawing there was a loss of protamine-1 combined with cryocapacitation-associated changes and apoptotic-like changes in buck spermatozoa. Spermatozoa deprotamination might be attributed to increased DNA fragmentation, resulting in compromised fertilizing capacity of buck spermatozoa.

Dynamics of HSPA1A and redox status in the spermatozoa and fluid from different segments of goat epididymis.

The present study was attempted to investigate the dynamics of HSPA1A and redox status in the spermatozoa and fluid of different segments of buck epididymis. Testes were collected from sexually mature and healthy bucks aged between 2 and 3 years. The fluid and spermatozoa from different segments (caput, corpus and cauda) were harvested for further processing and analysis. The concentration of HSPA1A in spermatozoa lysate and epididymal fluid and its relative mRNA expression in spermatozoa from different segments of epididymis were studied. The HSPA1A concentration in epididymal fluid was significantly (P < 0.01) higher in the corpus as compared with caput and cauda, whereas, its concentration and relative mRNA expression decreased significantly (P < 0.01) in the spermatozoa from caput to cauda. The activities of SOD, GR, GST, and concentrations of manoldialdehyde and ROS decreased significantly (P < 0.01) in the spermatozoa from caput to cauda. The glutathione concentration and GPx activity decreased significantly (P < 0.01) in the spermatozoa of cauda as compared with the corpus. The SOD activity and ROS concentration were significantly (P < 0.01) higher in corpus, and GR and GST activity were significantly (P < 0.01) higher in caput fluid as compared with corpus and cauda. It may be concluded that HSPA1A concentration and its relative mRNA expression in spermatozoa decreased progressively, and redox status was altered during transit from caput to cauda.

Functional insights into voltage gated proton channel (Hv1) in bull spermatozoa.

Acid extrusion and intracellular alkalisation are the key events during sperm capacitation and these are mediated through proton gated channels (Hv1). Role of Hv1 in regulating sperm motility, capacitation and acrosome reaction has been documented in human spermatozoa; but no such data is available in bull spermatozoa; therefore, the present study was undertaken in Hariana bull spermatozoa. Sixty four ejaculates were collected from four Hariana bulls to investigate the functional involvement of Hv1 in regulation of sperm motility, capacitation and acrosome reaction in bull spermatozoa. Immunoblotting revealed the presence of a single band of 31.8 kDa corresponding to Hv1 in Hariana bull spermatozoa and immunoflourescence confirmed the positive immune-reactivity at principal piece of spermatozoa for Hv1. Functional study was carried out using 200 µM 2-Guanidinobenzimidazole (2-Guanidinobenzimidazole,selective Hv1 blocker) and 1 mM zinc chloride (potent Hv1 blocker), and 0.3 µM Anandamide (AEA), an activator of Hv1. Blocking of Hv1 resulted in significant (P < 0.05) reduction in progressive sperm motility (PSM). Activation of Hv1 with AEA also resulted in significant (P < 0.05) reduction in PSM till 1 h and thereafter, the PSM was restored and reduction was almost similar to that in control. However, during activation and inactivation of Hv1, per cent spermatozoa showing hyperactive motility were found to be markedly increased (20-30%) compared to 10-20% in control. 2-Guanidinobenzimidazole, zinc and AEA treated spermatozoa revealed significant (P < 0.05) increase in B-pattern of spermatozoa indicating induction of capacitation. Downstream signalling of Hv1 activation or inactivation seemed to be mediated through cAMP and PKA pathways. Catsper channels were found to be intimately associated with Hv1 function in regulating sperm motility. Blocking as well as activation of Hv1 resulted in significant (P < 0.05) reduction in sperm livability, spermatozoa having intact membrane, intact acrosome, and high mitochondrial transmembrane potential (MTP). Our findings evidently suggest that Hv1 channels are present in bull spermatozoa and these regulate sperm functions like hypermotility, capacitation and acrosome reaction through complex interplay between different pathways involving cAMP, PKC, and Catsper. Further studies are required to find out the possible relationship between Hv1 channels and other channels in regulating spermatozoa functions.

Effect of sericin supplementation on heat shock protein 70 (HSP70) expression, redox status and post thaw semen quality in goat.

Cryopreservation results in substantial deterioration of heat shock protein 70 (HSP70) and ultra-structural changes in sperm organelles, resulting in a marked reduction in post-thaw semen quality. The present study was aimed to explicate the effect of sericin supplementation on expression profile of HSP70, redox status and post-thaw semen quality in Barbari goat. Five Barbari bucks were used to collect thirty semen ejaculates by using artificial vagina and each ejaculate was divided into three aliquots to which sericin was supplemented at 0% (Control), 0.25% (T1) and 0.50% (T2). Further, extended semen samples were equilibrated followed by their cryopreservation. Post-thaw semen characteristics, redox status of seminal plasma, enzyme leakage and HSP70 gene/protein expression in spermatozoa were assessed in all the groups. Per cent progressive motile spermatozoa, spermatozoa having intact plasma membrane (HOST + ve) and intact acrosomes in post-thaw spermatozoa were significantly (p < 0.01) higher in T1 and T2 as compared to control. A significant (p < 0.01) reduction in abnormal spermatozoa was found in T1 as compared to T2. Sericin supplementation significantly (p < 0.05) improved the antioxidative status (SOD, GST, CAT), reduced lipid peroxidation (MDA) and also prevented enzyme (ALT, LDH) leakage as compared to control samples. qRT-PCR results revealed that HSP70 mRNA expression was significantly (p < 0.01) upregulated in T1 and T2 group as compared to control. The positive effect of sericin on expression of HSP70 was further confirmed by immunoblotting followed by densitometry revealing higher expression in T1 and T2 compared to control. Inclusion of 0.25% w/v sericin in semen extender ameliorated the post-thaw semen quality by improving antioxidative status and minimizing the leakage of intracellular enzymes. Sericin supplementation had a beneficial effect on HSP70/HSP70 mRNA expression either by induction or by protection of HSP70/HSP70 mRNA as evident from the gene expression and immunoblotting studies.

Molecular characterization of voltage-gated potassium channel (Kv) and its importance in functional dynamics in bull spermatozoa.

Present study was undertaken to characterize the voltage gated potassium channel (Kv 1.1) in bull spermatozoa using sixty four ejaculates collected from four Hariana bulls. Functional characterization was undertaken using a selective blocker of Kv channel, 4-Aminopyridine (4-AP) while molecular presence of Kv on bull spermatozoa by immunoblotting and indirect immunofluorescence. Three sets of 100 µL diluted sperm samples namely-negative control (100 µL of sperm dilution medium (SDM) containing 10 × 106 cells), vehicle control (99 µL of SDM containing 10 × 106 cells, and DMSO- 1  µL) and 4-AP treatment group (99 µL of SDM containing 10 × 106 cells, and drug 1 µL 4-AP) were used in the study. Immunoblotting identified a single band of 56 kDa corresponding to Kv1.1 in Hariana bull spermatozoa. Immunolocalization showed the positive immunoreactivity at head, middle piece and principal piece of the spermatozoa for Kv 1.1. Blocking of Kv using 4-AP resulted in significant (p < 0.05) reduction in sperm progressive motility, per cent capacitated spermatozoa (B-pattern) and acrosome reacted (AR-pattern) spermatozoa, while significant (P < 0.05) increase in per cent swollen spermatozoa. Blocking of Kv channels resulted in significantly (P < 0.05) increased percentage of spermatozoa with lower mitochondrial transmembrane potential. Computer assisted semen analysis (CASA) of motion and kinematic parameters in 4-AP treated spermatozoa indicated reduction in sperm motion parameters like LIN, STR, VSL and VAP and higher ALH, VCL, and BCF indicating hyperactivity of spermatozoa. Based on our findings, it may be concluded that voltage-gated potassium channel (Kv) are present on bull spermatozoa and these are associated with functional dynamics of spermatozoa. However, based on our limited study, it is not possible to deduce that how these channels are associated with induction of hyperactivity. Therefore, further studies are warranted to unravel the mechanistic signaling pathways involved in Kv-mediated alterations in functional dynamics of spermatozoa.

Assessment of motion and kinematic characteristics of frozen-thawed Sirohi goat semen using computer-assisted semen analysis.

AIM: The aim was to determine the motion and kinematics characteristic of frozen-thawed spermatozoa in Sirohi goat using computer-assisted semen analysis. MATERIALS AND METHODS: A study was carried out in Sirohi buck. Semen collection was made biweekly from each buck with the help of artificial vagina. A total of 12 ejaculates were collected from two bucks (six ejaculates from each buck). Freshly collected semen was pooled and later evaluated. The pooled semen sample was extended with standard glycerolated egg yolk tris extender and later subjected to a process of cryopreservation. The motion and kinematic characteristics of spermatozoa were studied during freez-thawing process. RESULTS: Significantly (p<0.01) higher value of live percent, hypo-osmotic swelling test, and acrosomal integrity were recorded in neat semen followed by diluted and frozen thaw semen. The proportion of spermatozoa showing slow progression were the highest in the neat and diluted semen followed by rapid and non-progressively motile, while a reverse pattern was observed in the frozen thaw semen where the proportion of non-progressively motile spermatozoa were significantly (p<0.01) higher followed by slow and rapid progression. CONCLUSION: This study showed that the best results for motion, vitality, plasma membrane integrity, and acrosome status were obtained in the neat semen followed by diluted and frozen thaw semen. Further, the process of cryopreservation results in a shift of motility from slow to non-progressive in the post-thaw semen with a significant decrease in the path velocities when compared to neat and diluted semen. Hence, it can be concluded that freezing-thawing process reduces the motility and kinematic characters spermatozoa and may be an important factor affecting the fertilizing ability of spermatozoa resulting in poor conception rate after insemination in goats.