PubMed Health⌕ Search

Biomedical subjects

T L Blanchard

Publications and source records attributed to T L Blanchard.

At least 19 recordsLinked to original sources
PubMed2005

Relationship of seminal plasma level and extender type to sperm motility and DNA integrity.

The relationship between seminal plasma level (0, 10, or 20%) and extender type [Kenney type (EZ-Mixin-CST) or Kenney-modified Tyrodes-KMT] to the susceptibility of sperm DNA to denaturation and sperm motility measures were investigated in cooled (5 degrees C) stallion sperm. Three ejaculates from each of three fertile stallions were collected in an artificial vagina and processed as follows: diluted one part uncentrifuged semen with four parts of extender to a final concentration of 20% seminal plasma in either CST or KMT (20% CST; 20% KMT); diluted to a final concentration of 25 million sperm/mL in either CST or KMT (10% CST; 10% KMT); centrifuged to remove virtually all seminal plasma and resuspended in either CST or KMT (0% CST-Cent; 0% KMT-Cent); centrifuged semen to remove virtually all seminal plasma and resuspended with previously filtered seminal plasma from the same stallion in either CST or KMT to a final concentration of 20% seminal plasma (20% CST-Cent; 20% KMT-Cent). Sperm motion characteristics were determined by CASA and DNA integrity (%COMP, percent of cells outside the main population) evaluated by the Sperm Chromatin Structure Assay prior to cooling, and after 24 and 48 h cooled-storage at 5 degrees C. After 48 h of storage at 5 degrees C, extenders with 0% seminal plasma (0% CST-Cent, 0% KMT-Cent) maintained highest quality DNA (P < 0.05), but 0% KMT-Cent maintained higher velocity measures (P < 0.05) than 0% CST-Cent. Total sperm motility was highest (P < 0.05) in 0% CST-Cent, 0% KMT-Cent, 10% CST, 20% CST-Cent, and 20% CST compared to the other treatment groups. Progressive sperm motility was highest (P < 0.05) after 48 h of storage in the treatment with 10% seminal plasma in Kenney extender (10% CST), despite a reduction in DNA integrity. Regardless of extender type, addition of 20% seminal plasma following centrifugation resulted in almost a two-fold increase in %COMP(alpha t), even though one of the treatments (20% CST-Cent) maintained total and progressive motility similar to treatments with no seminal plasma, suggesting that sperm motility and DNA integrity may respond independently to environmental conditions. Overall, better quality sperm features (motility and DNA) were maintained in sperm from which seminal plasma was removed followed by resuspension in either Kenney extender or modified Kenney Tyrodes-type extender.

Animals↗
PubMed2004

Effects of a new injectable short-term release deslorelin in foal-heat mares.

Mares treated with subcutaneous deslorelin implants on the first postpartum estrus early in the breeding season had significant reductions in the number of large follicles at early pregnancy examinations and delayed return to estrus (in mares that failed to become pregnant); these adverse effects were attributed to a prolonged release of the drug from the implant. In 2003, an injectable short-term release (<24 h) deslorelin product became available. The objective of this study was to determine if this product would hasten ovulation in early foaling first postpartum estrus mares without reducing the number of large follicles at early pregnancy examination (14-15 days postovulation). Beginning 5-6 days postpartum, first postpartum estrus (foal-heat) mares were teased daily and examined thrice weekly (Tuesday, Thursday and Saturday) by transrectal ultrasonography. Mares in estrus with a follicle > or = 34 mm diameter on Tuesdays or Thursdays were alternately assigned to: Treatment 1, n = 17; 1.5 mg injectable short-term release deslorelin, or Treatment 2, n = 16; Control (no treatment). The schedule allowed accurate determination of the number of mares ovulating within 2 days of treatment (i.e., ovulations detected on Thursday or Saturday). Mares were mated on the day of treatment and at 2-day intervals until either ovulation was confirmed or until behavioral estrus ceased. Transrectal ultrasonography was done 14-15 days after ovulation to assess ovarian follicles and pregnancy status. Fewer covers were required and more mares ovulated within 2 days of treatment in deslorelin-treated versus Control mares (P < 0.01). Pregnancy rates were normal (69%) in deslorelin-treated mares. The number of large follicles 14-15 days after ovulation did not differ between deslorelin-treated and Control mares (P > 0.10), suggesting follicular suppression did not occur with this formulation of deslorelin.

Animals↗
PubMed2003

Relationship between stallion sperm motility and viability as detected by two fluorescence staining techniques using flow cytometry.

Relationships between sperm motility parameters and viability were evaluated using two fluorescent staining techniques in fresh extended semen (fresh and after 24 h storage at 5 degrees C) that had various concentrations of dead sperm added to simulate different levels of viable and nonviable sperm. Both protocols incorporated SYBR-14 and propidium iodide (PI) while the second protocol added the mitochondrial probe JC-1. The relationship between total sperm motility and percent viable sperm was high between staining protocols (r = 0.98). Time (0 h versus 24 h, P<0.0001) and treatment (0, 10, 25, 50, and 75% nonviable sperm, P<0.0001) affected percent total sperm motility and percent viable sperm for both staining protocols. Actual percent viable sperm for each time and treatment did not differ from expected values.

Animals↗
PubMed2003

Effects of dead spermatozoa on motion characteristics and membrane integrity of live spermatozoa in fresh and cooled-stored equine semen.

The aim of this study was to determine if dead spermatozoa reduced motility or membrane integrity of live spermatozoa in fresh and cooled-stored equine semen. Three ejaculates from each of three stallions were centrifuged and virtually all seminal plasma was removed. Spermatozoa were resuspended to 25 x 10(6) spermatozoa/ml with EZ-Mixin CST extender and 10% autologous seminal plasma, then divided into aliquots to which 0 (control), 10, 25, 50, or 75% (v/v) dead spermatozoa were added. Dead spermatozoa preparations contained 25 x 10(6) spermatozoa/ml and 10% seminal plasma from pooled ejaculates of the three stallions, in EZ-Mixin CST extender. Spermatozoa were killed in the pooled ejaculates by repeated freezing and thawing, then stored at -20 degrees C until warmed to 37 degrees C and mixed with aliquots of fresh spermatozoa to be cooled and stored in an Equitainer for 24h. Motion characteristics (% total motility (MOT), % progressive motility (PMOT), and mean curvilinear velocity (VCL)) for fresh and 24h cooled samples were determined using a computerized spermatozoal motion analyzer. The presence of up to 75% dead spermatozoa did not adversely affect MOT or PMOT of live spermatozoa in either fresh or cooled-stored semen. However, VCL and the percentage of membrane-intact spermatozoa were reduced compared to control samples when 75% (v/v) dead spermatozoa were added. Membrane integrity, as assessed by staining with carboxyfluoresein diacetate-propidium iodide, was highly correlated (r>0.8; P<0.001) with MOT and PMOT in both fresh and cooled-stored semen samples. Results of this study have application to the processing of both cooled and frozen equine semen.

Animals↗
PubMed2002

Insulin-like growth factor-I and insulin-like growth factor binding protein-2 and -5 in equine seminal plasma: association with sperm characteristics and fertility.

The objectives of this study were 1) to determine whether insulin-like growth factor-I (IGF-I) and insulin-like growth factor binding proteins (IGFBPs) were present in seminal plasma of stallions; 2) to compare semen parameters (IGF proteins, sperm numbers, morphology, and motility) from stallions at sexual rest (SR) and when sexually active (SA); 3) to compare semen parameters between stallions with high and low seminal plasma IGF-I concentrations; and 4) to examine the relationship between seminal plasma IGF-I concentrations and fertility parameters of stallions. Ejaculates were collected from stallions at SR (n = 51) and SA (n = 46). Concentrations of IGF-I and IGFBP-2 in seminal plasma samples were determined by radioimmunoassay. Presence of IGFBPs in equine seminal plasma was verified using immunoprecipitation and Western ligand blot procedures. IGF-I, IGFBP-2, and IGFBP-5 were present in equine seminal plasma. Concentrations of IGF-I, IGF-I/protein, total IGF-I, IGFBP-2, IGFBP-2/protein, and total IGFBP-2 were not significantly different (P > or = 0.13) in seminal plasma between stallions at either SR or SA. At SR, stallions with higher seminal plasma IGF-I had more total IGFBP-2 per ejaculate (P < 0.01), more morphologically normal sperm (P = 0.05), and higher first-cycle pregnancy rates (P = 0.02). At SA, stallions with higher seminal plasma IGF-I had fewer cycles per pregnancy (P = 0.02). An association of seminal plasma IGF-I concentration with sperm motility, sperm morphology, and pregnancy rates in bred mares suggests that IGF-I may play a role in sperm function.

Animals↗
PubMed2001

Advances in cooled semen technologies: seminal plasma and semen extender.

This study evaluated motility and fertility of uncentrifuged and centrifuged equine semen following dilution in a skim milk-glucose extender with or without supplemental Tyrode's medium. In addition, the effect of seminal plasma addition to each extender was evaluated. For Experiment 1, motility of 48h cooled, stored spermatozoa was evaluated following eight dilution treatments: uncentrifuged and diluted 1:4 (v/v) in skim milk-glucose extender (EZ Mixin CSTJ; CST-1:4) or in CST supplemented 65:35 (v/v) with modified Tyrode's medium (KMT-1:4); uncentrifuged and diluted to 25x10(6) spermatozoa/ml in CST (CST-1:9) or in KMT (KMT-1:9); centrifuged and diluted in CST with 0% seminal plasma (CST-0) or 20% seminal plasma (CST-20) or centrifuged and diluted in KMT containing 0% seminal plasma (KMT-0) or in KMT containing 20% seminal plasma (KMT-20). Sperm motility parameters evaluated included percentage of total motile sperm (% TMOT), percentage of progressively motile sperm (% PMOT), curvilinear velocity (VCL) and straight-line velocity (VSL). Mean % PMOT was lower (P<0.05) for spermatozoa extended in CST-1:4 compared to CST-1:9, whereas, all motility parameters were reduced (P<0.05) in KMT-1:4 compared to KMT-1:9. Spermatozoa extended in CST-1:4 had greater % TMOT, % PMOT and VSL (P<0.05) than in KMT-1:4. Spermatozoa extended in CST-1:9 had greater (P<0.05) % PMOT than in KMT-1:9, however, VCL was greater (P<0.05) in KMT-1:9. Mean VCL and VSL were lower (P<0.05) for spermatozoa extended in CST-0 compared with CST-20, whereas, spermatozoa extended in KMT-0 had greater (P<0.05) % TMOT, % PMOT and VSL compared to spermatozoa extended in KMT-20. Mean % TMOT and % PMOT were greater (P<0.05) in CST-20 compared to KMT-20, however, KMT-0 increased (P<0.05) velocity measures (VCL and VSL) compared to CST-0. In Experiment 2, fertility of centrifuged spermatozoa diluted in either CST-20 or KMT-0 was similar (P>0.05). We conclude that modified Tyrode's medium was not detrimental to establishment of pregnancy. Use of modified Tyrode's medium may improve spermatozoal motility and pregnancy rates for cooled transport of semen from stallions in which all seminal plasma must be removed because of suspected toxic effects of seminal plasma on spermatozoal viability, however, Tyrode's medium may be detrimental to sperm motility when seminal plasma is present.

Animals↗
PubMed2001

Mares with delayed uterine clearance have an intrinsic defect in myometrial function.

Persistent, postmating endometritis affects approximately 15% of mares and results in reduced fertility and sizable economic losses to the horse-breeding industry. Mares that are susceptible to postmating endometritis have delayed uterine clearance associated with reduced uterine contractility. Unfortunately, the mechanism for reduced uterine contractility remains an enigma. The present study examined the hypothesis that mares with delayed uterine clearance have an intrinsic contractile defect of the myometrium. Myometrial contractility was evaluated in vitro by measuring isometric tension generated by longitudinal and circular uterine muscle strips in response to KCl, oxytocin, and prostaglandin F(2alpha) (PGF(2alpha)) for young nulliparous mares, older reproductively normal mares, and older mares with delayed uterine clearance. In addition, intracellular Ca(2+) regulation was evaluated using laser cytometry to measure oxytocin-stimulated intracellular Ca(2+) transients of myometrial cells loaded with a Ca(2+)-sensitive fluorescent dye, fluo-4. For all contractile agonists, myometrium from mares with delayed uterine clearance failed to generate as much tension as myometrium from older normal mares. Oxytocin-stimulated intracellular Ca(2+) transients were similar for myometrial cells from mares with delayed uterine clearance and from older normal mares, suggesting that the contractile defect did not result from altered regulation of intracellular Ca(2+) concentration. Furthermore, no apparent age-dependent decline was observed in myometrial contractility; KCl-depolarized and oxytocin-stimulated longitudinal myometrium from young normal mares and older normal mares generated similar responses. However, circular myometrium from young normal mares failed to generate as much tension as myometrium from older normal mares when stimulated with oxytocin or PGF(2alpha), suggesting possible age-related alterations in receptor-second messenger signaling mechanisms downstream of intracellular Ca(2+) release. In summary, for mares with delayed uterine clearance, an intrinsic contractile defect of the myometrium may contribute to reduced uterine contractility following breeding.

Aniline Compounds↗
PubMed2000

Techniques for evaluating selected reproductive disorders of stallions.

Numerous techniques may be used for evaluation of the different reproductive disorders of the stallion. Approaches may vary from real-time ultrasonography and biopsy for evaluating testicular tumors to use of special assays for evaluating sperm or plasma for presence of antisperm antibodies. This communication addresses techniques used to evaluate five relatively uncommon, but perplexing, disorders of breeding stallions: (1) seminal vesiculitis, (2) hemospermia associated with idiopathic urethral defects, (3) acrosomal dysfunction, (4) abnormal spermatozoal chromatin, and (5) azoospermia.

Animals↗
PubMed2000

Effects of transport container and ambient storage temperature on motion characteristics of equine spermatozoa.

This study was conducted to compare the cooling rates and storage temperatures within equine semen transport containers exposed to different ambient temperatures, and to evaluate the ability of these containers to preserve spermatozoal motility following 24 h of storage under these conditions. In Experiment 1, nonfat dried milk solids, glucose, sucrose, equine semen extender was divided into seven 40-mL aliquots and loaded into seven different semen transport containers: Equitainer I, Equitainer II, Equitainer III, ExpectaFoal, Bio-Flite, Lane STS, and Equine Express. After containers were loaded, they were subjected to one of three ambient storage temperatures: 1) 22 degrees C for 72 h, 2) -20 degrees C for 6 h followed by 22 degrees C for 66 h, or 3) 37 degrees C for 72 h. Cooling rates and storage temperatures of semen extender in each container were monitored with thermocouples and a chart recorder. In Experiment 2, semen from each of three stallions (3 ejaculates per stallion) was diluted to 25 x 10(6) spermatozoa/mL with semen extender, divided into 40 mL aliquots and loaded into transport containers as in Experiment I. Containers were subjected to one of three ambient storage conditions: 1) 22 degrees C for 24 h, 2) -20 degrees C for 6 h, followed by 22 degrees C for 18 h, or 3) 37 degrees C for 24 h. After 24 h of storage, spermatozoal motion characteristics (percentage of motile spermatozoa; MOT, percentage of progressively motile spermatozoa; PMOT, and mean curvilinear velocity; VCL) were evaluated using a computerized spermatozoal motion analyzer. Significant interactions were detected among storage conditions and semen transport containers for the majority of the temperature endpoints measured. When exposed to temporary ambient freezing conditions, the lowest temperatures attained by samples in containers ranged from -2.8 to 0.8 degrees C. Lowest temperature samples attained was not correlated (P > 0.05) with spermatozoal motility under any ambient condition. However, time below 4 degrees C was highly correlated (P < 0.05) with a reduction in spermatozoal motility. Mean cooling rates from 20 degrees C to 8 degrees C did not correlate with spermatozoal motility, except when containers were exposed to temporary freezing conditions. No container cooled samples below 6 degrees C in 22 degrees C or 37 degrees C environments except for the ExpectaFoal, in which samples fell below 4 degrees C under all ambient conditions. Ambient temperature affected MOT, PMOT and VCL of semen stored in all containers (P < 0.05) except for the Equitainer II in which motion characteristics remained high and were similar among all ambient temperatures (P > 0.05). Results suggest that stallion semen may be able to tolerate a wider range of cooling rates and storage temperatures than previously considered safe.

Animals↗
PubMed1999

Estrous cycle characteristics and response to estrus synchronization in mammoth asses (Equus asinus americanus).

Breeding records from a herd of mammoth asses (Equus asinus americanus) maintained on pasture in southeast Texas from 1990 to 1998 were reviewed. Jennies were pasture or hand mated, and estrus was either observed while the jennies were on pasture or when exposed to a jack after being penned. Eighty-one estrus periods and 43 diestrus intervals were recorded in 33 jennies over 4 seasons of the year (January-March, April-June, July-September, and October-December). Estrous cycle length and the duration of estrus were similar among seasons. Over all seasons, estrous cycle length was 23.3 +/- 2.6 d, duration of estrus was 5.9 +/- 2.1 d, and diestrus length was 17.4 +/- 2.6 d (mean +/- SD). During these same 9 yr, 58 injections of PGF2 alpha (5 mg, i.m.) were administered to 38 jennies without regard to stage of estrous cycle. Seventy-six percent (44/58) of the jennies showed signs of estrus after PGF2 alpha treatment, with an interval to estrus of 4.4 +/- 1.6 d and a duration of estrus of 5.6 +/- 1.7 d. Two estrus synchronization schemes were also assessed. Trial 1 was performed in October to November 1996, and Trial 2 was performed in February to March 1998. In Trial 1 (Group PE + PGF, n = 10), each jenny was injected intramuscularly once daily for 10 d with 150 mg progesterone and 10 mg estradiol-17 beta in sesame oil, and PGF2 alpha (10 mg) was injected intramuscularly on the last day of treatment. In Trial 2 (Group PGF-2X, n = 11), each jenny was injected intramuscularly twice, 16 d apart, with 10 mg PGF2 alpha. All Group PE + PGF jennies responded to treatment. One jenny in Group PGF-2X did not respond to either injection of PGF2 alpha, while 2 jennies responded to the first but not the second PGF2 alpha injection (8 of 11 jennies returned to estrus and ovulated after the second PGF2 alpha injection). Duration of estrus was 6.8 +/- 1.9 d for Group PE + PGF and 7.1 +/- 1.8 d for Group PGF-2X jennies. Interval to estrus and interval to ovulation following the last treatment were 9.0 +/- 0.9 d and 14.5 +/- 1.7 d, respectively, in Group PE + PGF jennies, and 4.5 +/- 0.9 d and 10.4 +/- 1.8 d, respectively, for Group PGF-2X jennies. In summary, estrous cycle characteristics of mammoth asses are similar to those reported for standard jennies, and estrus synchronization schemes used in horses are effective in mammoth asses.

Animals↗
PubMed1998

Fertility comparison between breeding at 24 hours or at 24 and 48 hours after collection with cooled equine semen.

It has become a common practice in the equine breeding industry to send 2 insemination doses for breeding with transported cooled semen, one to be used for the initial insemination upon arrival, and the other to be held a second insemination the next day. One fertile stallion and 36 fertile mares were used to determine if breeding once with 1 dose of semen cooled for 24 h would improve fertility compared with breeding twice, 1 d apart, with half the dose of semen cooled for 24 h on the first day of breeding and half cooled for 48 h on the second day of breeding. Mares were given two intramuscular injections of 10 mg PGF2 alpha 14 d apart. Following the second injection, mares were teased with a stallion and their ovaries were scanned by transrectal ultrasonography daily. When a dominant follicle (> 35 mm diameter) was detected, 1500 units hCG were injected intravenously, and the mares were inseminated. Semen was collected in advance of anticipated breeding, mixed in nonfat dry milk solids-glucose extender to a concentration of 25 million sperm/mL, and placed in 2 commercial cooling containers for 24 or 48 h of storage prior to breeding. Mares were randomly assigned to 1 of 2 insemination treatment groups: 1) Group T1 (n = 18), in which mares were inseminated on the day of hCG injection with 500 million spermatozoa cooled for 24 h, or 2) Group T2 (n = 18), in which mares were inseminated on the day of hCG injection with 250 million spermatozoa cooled for 24 h, and again on the following day with 250 million spermatozoa cooled for 48 h. Pregnancy status was confirmed by transrectal ultrasonographic examination at 14 and 16 d after ovulation. Pregnancy rates were the same for both insemination treatment groups (12/18; 67%). There was no advantage to holding half of the insemination dose for rebreeding on the following day.

Animals↗
PubMed1998

Bacteriology of preserved stallion semen and antibiotics in semen extenders.

Three experiments were conducted to evaluate the effects of different antibiotics in a milk-glucose semen extender on motility of equine sperm and elimination of bacteria following storage of extended semen in vitro. In Experiment 1, 7 antibiotics were compared: amikacin, gentamicin, streptomycin, potassium penicillin, sodium penicillin, ticarcillin, and polymixin B. In Experiment 2, 3 antibiotic treatments were compared: potassium penicillin G, amikacin, or a combination of potassium penicillin G and amikacin. In Experiment 3, 3 antibiotic treatments were compared: potassium penicillin G-amikacin, ceptiofur, and a combination of ticarcillin and clavulanic acid (Timentin). Control treatments (antibiotic-free extender) were included in each experiment. Six motility variables were evaluated: percentage of motile sperm; percentage of progressively-motile sperm; percentage of rapidly-motile sperm; mean curvilinear velocity; mean average path velocity; and mean straight-line velocity. In Experiment 1, mean percentages of motile, progressively motile and rapidly motile sperm were lower (P < 0.05) in semen exposed to polymixin B then in other treatments. Mean average-path velocity of sperm in extender containing polymixin B was lower (P < 0.05) than that of all other treatments, with exception of control or ticarcillin. Mean straight-line velocity of sperm in extender containing polymixin B was lower (P < 0.05) than that of all other treatments, with exception of control, streptomycin or ticarcillin. Semen samples containing gentamicin, amikacin, streptomycin, or potassium penicillin were more effective (P < 0.05) at eliminating bacterial growth than those samples containing polymixin B. Semen samples containing gentamicin were also more effective (P < 0.05) at eliminating bacterial growth than those samples containing ticarcillin or sodium penicillin. In Experiment 2, mean percentage of rapidly-motile sperm, and mean curvilinear, average-path, and straight-line velocities were greater (P < 0.05) for potassium penicillin-amikacin than values for all other treatments. In 2 of 3 stallions, an effect of treatment on percentage of motile sperm was detected (P < 0.05). For one stallion, mean motility of potassium penicillin-amikacin was greater (P < 0.05) than that of all other treatment groups. For another stallion, mean motility of the control was lower (P < 0.05) than that of the other treatments. Following storage, potassium penicillin (16/18 [89%]) or potassium penicillin-amikacin (17/19 [94%]) were more effective (P < 0.05) at controlling aerobic and anaerobic bacterial isolates in semen specimens than was amikacin (10/18 [56%]). In Experiment 3, a difference among treatment groups for motility variables was not detected (P < 0.05). No bacterial growth was recovered in antibiotic-treated semen, with exception of Micrococcus sp. (2 colonies) which were isolated from one semen specimen treated with ceptiofur.

Amikacin↗
PubMed1997

Three methods of oxytocin-induced parturition and their effects of foals.

OBJECTIVE: To compare effects of 3 oxytocin-based induction techniques on fetal and neonatal foals. DESIGN: Prospective randomized controlled trial. ANIMALS: 16 pregnant mares. PROCEDURE: Parturition was induced in mares by use of 3 treatments: group 1, 75 U of oxytocin, IM; group 2, 15 U of oxytocin, IM, q 15 minutes, for a maximum of 75 U; group 3, 75 U of oxytocin in 1 L of 0.9% NaCl solution IV (1 U/min), for a maximum of 75 U. Blood gas values and indices of vitality were measured in foals, and variables describing parturition were measured in mares. RESULTS: Group-3 mares had a shorter interval from administration of oxytocin to rupture of the chorioallantois (OTCA) than group-2 mares. More foals were abnormal when the interval from oxytocin administration to delivery of the foal (OTDE) was > or = 60 minutes. Arterial blood gas values, measurements of vitality, and plasma cortisol concentrations did not differ among foals in various treatment groups. Increased interval for OTCA and OTDE resulted in higher neonatal PaCO2, and a longer interval for OTCA resulted in lower arterial pH. Time required for birth was shorter in mares with a dilated cervix. More abnormal foals than normal foals were delivered after premature placental separation or dystocia. Abnormal foals took longer to stand and suckle than normal foals. Interval from delivery to suckling was positively correlated with OTCA, OTDE, and PaCO2. CLINICAL IMPLICATIONS: Method of oxytocin-induced parturition did not impact neonatal outcome. Interval from induction until parturition, degree of cervical dilatation, and intrapartum complications influenced induction success.

Animals↗
PubMed1997

Determination of daily sperm production in stallion testes by enumeration of germ cells in homogenates.

Thirty adult stallion testes were selected with high (n = 15) and low (n = 15) Daily Sperm Production (DSP)/testis. Parenchymal samples were prepared for morphometric analysis, and the numbers of germ cells and Sertoli cells were determined. Testicular samples were homogenized, and germ cells and Sertoli cells were enumerated using phase contrast microscopy. Numbers of germ cells and Sertoli cells and potential DSP during spermatogenesis were determined. Significant correlations existed between morphometric and homogenate determinations of number per testis of preleptotene, leptotene plus zygotene primary spermatocytes (r = 0.58; P < 0.001), pachytene plus diplotene primary spermatocytes (r = 0.67; P < 0.0001), all primary spermatocytes (r = 0.67; P < 0.0001), round spermatids (r = 0.72; P < 0.0001), and Sertoli cells (r = 0.70; P < 0.0001). Significant correlations (P < 0.0001) existed between morphometric and homogenate determination of DSP/testis based on preleptotene, leptotene plus zygotene primary spermatocytes (r = 0.78), pachytene plus diplotene primary spermatocytes (r = 0.88), and round spermatids (r = 0.85). Using morphometric determination as the standard, the sensitivity (i.e., ability to detect low DSP/testis) and specificity (i.e., ability to detect high DSP/testis) by homogenate enumeration of germ cells was 81 and 93% for round spermatids, 100 and 24% for pachytene plus diplotene primary spermatocytes, and 67 and 87% for preleptotene, leptotene plus zygotene primary spermatocytes, respectively. Enumeration of primary spermatocytes in homogenates was less accurate than enumeration of round or elongated spermatids. Enumeration of round and elongated spermatids in homogenates was a rapid and useful method for determining DSP in horses, and it may prove to be a useful technique for quantitating potential DSP from testicular biopsies.

Journal Article↗
PubMed1997

Increased germ cell degeneration and reduced germ cell:Sertoli cell ratio in stallions with low sperm production.

To determine the relationship between germ cell degeneration or germ cell:Sertoli cell ratio and daily sperm production, testes were obtained during the months of May to July (breeding season) and November to January (nonbreeding season) from adult (4 to 20-yr-old) stallions with either high (n = 15) or low (n = 15) sperm production. Serum was assayed for concentrations of LH, FSH and testosterone. Testes were assayed for testosterone content and for the number of elongated spermatids, after which parenchymal samples were prepared for histologic assessment. Using morphometric procedures, the types and numbers of spermatogonia, germ cells and Sertoli cells were determined. High sperm producing stallions had greater serum testosterone concentration, total intratesticular testosterone content, testicular parenchymal weight, seminiferous epithelial height, diameter of seminiferous tubules, numbers of A and B spermatogonia per testis, number of Sertoli cells per testis, and number of B spermatogonia, late primary spermatocytes, round spermatids and elongated spermatids per Sertoli cell than low sperm producing stallions (P < 0.05). The number of germ cells (total number of all spermatocytes and spermatids in Stage VIII tubules) accommodated by Sertoli cells was reduced in low sperm producing stallions (18.6 +/- 1.3 germ cells/Sertoli cell) compared with that of high sperm producing stallions (25.4 +/- 1.3 germ cells/Sertoli cell; P < 0.001). The conversion from (yield between) early to late primary spermatocytes and round to elongated spermatids was less efficient for the low sperm producing stallions (P < 0.05). Increased germ cell degeneration during early meiosis and spermiogenesis and reduced germ cell:Sertoli cell ratio was associated with low daily sperm production. These findings can be explained either by a compromised ability of the Sertoli cells to support germ cell division and/or maturation or the presence of defects in germ cells that predisposed them to degeneration.

Journal Article↗
PubMed1997

Predicting end-of-test semen quality in bulls prior to performance testing.

The first objective of this study was to determine if serum concentrations of specific hormones (testosterone, progesterone and androstenedione) in bulls at the start of performance testing could predict semen quality at the end-of-test when used in a multivariate model. The second objective was to evaluate other clinical measurements (breed, age, body weight, hip height and scrotal circumference) for predicting end-of-test semen quality. End-of-test semen quality was related to steroid concentrations and several pre-testing measurements, including age, body weight, hip height and scrotal circumference (SC). Combining the 3 steroid concentrations into a predictive test had a sensitivity of 0.6 and specificity of 0.5 at its most accurate point. The repeatability of the test result was extremely low (r(2) = 0.16; P < 0.05). In multivariate analyses, breed and start-of-test SC remained significant predictors of end-of-test semen quality (P < 0.05) while the other variables were nonsignificant (P > 0.1), suggesting that start-of-test SC was the most accurate predictor of end-of-test semen quality. Removing bulls at the start-of-test that had scrotal measurements of less than 20 cm, 24 cm, 28 cm or 32 cm resulted in sensitivities and specificities of 0.19, 0.94; 0.41, 0.81; 0.64, 0.56; and 0.94, 0.12, respectively. No cut-point had both adequate sensitivity and specificity. Because clinical tests were correlated, combining the tests to improve accuracy was not justified.

Journal Article↗
PubMed1997

Comparison of three containers used for the transport of cooled stallion semen.

Three containers commonly used to transport cooled equine semen (Equitainer, ExpectaFoal and a Swedish-designed semen-transport container, previously called the Salsbro Box and now called Equine Express) were compared, using four ejaculates from each of three stallions. Each ejaculate was diluted to a spermatozoal concentration of 25 x 10(6)/ml with a nonfat dry milk-glucose extender containing amikacin sulfate (1 mg/ml) and potassium penicillin G (1000 units/ml). Extended semen was divided into three 40-ml aliquots for placement in each of the three semen-transport containers. The extended semen was stored in the containers for 24 h prior to analysis. Stored semen was warmed for 15 min at 37 degrees C, then video records of sperm motility were obtained for evaluation using a Hamilton-Thorne motility analyzer equipped with a stage warmer set at 37 degrees C. The temperature of 40-ml aliquots of semen extender stored in each container was also measured for 60 h using a copper-constantan thermocouple placed in the center of the stored samples. Intervals from onset of storage until sample temperature exceeded 10 degrees C during the warming phase were 27.5, 33.5 and 53 h, for the Expecta-Foal, Equine Express and Equitainer, respectively. Semen extender stored in the Equitainer compared most favorably to ideal cooling rates and storage temperatures published previously. Following a 24-h storage period, the mean percentages of motile, progressively motile, and rapidly motile spermatozoa, as well as the mean spermatozoal curvilinear velocity were similar (P > 0.05) among the three containers.

Journal Article↗