Chromatin structural changes in sperm after scrotal insulation of Holstein bulls.

The reported effects on semen quality ascribed to testicular heat stress generally relate to traits impacting sperm transport and fertilizing ability but not to the genetic material contained by the sperm. To characterize the effects of testicular heat stress on sperm chromatin, susceptibility of DNA in sperm nuclear chromatin to in situ acid denaturation was measured by flow cytometry after staining with acridine orange using the sperm chromatin structure assay (SCSA). Semen was collected from Holstein bulls at 3-day intervals, before and after 48-hour scrotal insulation, until the morphologically abnormal sperm content in raw semen exceeded 50%. After cryopreservation in egg yolk-citrate extender, semen was thawed and sampled during incubation in vitro at 38.5 degrees C. Overall, SCSA results showed that chromatin susceptibility to denaturation was increased for sperm collected post- vs. preinsulation and was more pronounced for sperm presumably in the testes during insulation than for those sperm presumably in the epididymides. Increased susceptibility was detected as early as the first collection postinsulation; however, chromatin of sperm presumably in the proximal epididymis during insulation did not appear to have been detrimentally affected. Chromatin susceptibility to denaturation increased with increased incubation time in vitro, but the rate of change in susceptibility during incubation did not differ among pre- vs. postinsulation specimens. We conclude that elevated scrotal temperatures adversely affect both epididymal and testicular sperm by reducing sperm chromatin stability. The effects of heat stress on the chromatin of epididymal sperm were more subtle than those exhibited by testicular sperm but detectable within close proximity to the heat stress event.

Accessory sperm: their importance to fertility and embryo quality, and attempts to alter their numbers in artificially inseminated cattle.

Accessory sperm number and its relationship to fertilization and embryo quality was evaluated in cattle after nonsurgical recovery of ova or embryos 6 d after insemination. Efforts to alter accessory sperm number per ovum included 1) blockage of retrograde sperm loss at insemination using a modified insemination device, 2) elevated sperm number per inseminate (40 x 10(6) vs 20 x 10(6], and 3) alteration in semen quality (percentage of viable and morphologically normal sperm in the inseminate). None of these efforts affected accessory sperm number per ovum or embryo. However, blockage of retrograde semen flow for 3 h or use of semen of below-average quality (decreased percentage of viable and morphologically normal sperm) resulted in significant decreases in number of viable embryos and increases in number of degenerate embryos and unfertilized ova compared with conventional insemination (P less than .03) and use of semen with an average percentage of viable and morphologically normal sperm (P less than .06). Number of accessory sperm per embryo or ovum was positively related to fertilization and embryo quality (P less than .05). Mean accessory sperm +/- SD and the median value (in parentheses) for unfertilized ova, degenerate embryos, and embryos classified fair to poor and excellent to good were, respectively, .3 +/- .8 (0), 5.4 +/- 8.9 (1.0), 15.8 +/- 28.6 (3.5), and 16.9 +/- 29.5 (5.0). We conclude that efforts to improve accessory sperm numbers per embryo or ovum failed and that high variation and skewness of accessory sperm toward 0 may make median values more meaningful than means.(ABSTRACT TRUNCATED AT 250 WORDS)

Effects of scrotal insulation on viability characteristics of cryopreserved bovine semen.

The effect of a 48-h scrotal insulation on spermatozoal viability (motility and acrosomal integrity), before and after semen cryopreservation, was studied in six young Holstein bulls whose semen was collected twice in succession at 3-d intervals. Motility and acrosomal integrity were measured before and after incubation of semen at 37 degrees C for 3 h. For assessment of results, collection days were grouped: period 1 (control) = d -6, -3, and 0, where d 0 = initiation of scrotal insulation after semen collection; period 2 = d 3, 6, and 9 (sperm presumed in the epididymis or rete testis during scrotal insulation); period 3 = d 12, 15, ... 39 (sperm presumed in spermatogenesis during scrotal insulation). Semen was cryopreserved each collection day until morphologically abnormal cells exceeded 50% of the ejaculate (d 12 to 21). Semen viability before and after freezing was lower in period 3 than in period 1 (P less than .05). These differences coincided with the appearance in period 3 of abnormal sperm morphology and depressed undiluted semen motility, which began on d 12 (P less than .01). Semen collected during period 2 that was extended but unfrozen did not differ from that collected during period 1 in morphology or viability. However, for frozen semen, period 2 was significantly poorer than period 1 for both viability measurements, but only after incubation for 3 h at 37 degrees C postthaw (P less than .05). We conclude that epididymal sperm are adversely affected by elevated testicular temperatures, as noted by their decreased ability to maintain motility and acrosomal integrity following cryopreservation.