Three dairy heifers were examined during three consecutive oestrous cycles (control period, CP). Subsequently, the animals were subjected to 4 and then 5 weeks of twice-weekly ovum pick-up (OPU) (FPP1 and FPP2, respectively) with a recovery period (RP) of two consecutive oestrous cycles between FPP1 and FPP2. After FPP2, the animals were slaughtered and the ovaries were macroscopically examined. Throughout, ovarian activity was monitored by transrectal ultrasonography and concentrations of plasma progesterone. During CP, all the heifers showed normal cyclicity. During FPPs, the heifers occasionally presented oestrous activity. Corpus luteum (CL)-like structures developed from punctured follicles with diameters and life-spans varying from smaller and shorter than those in the CP (P > 0.05) to equal to those in the CP. There was a tendency for a lower number of emerging and punctured follicles in the presence of a CL-like structure. Subsequently to FPP1, all heifers regained normal cyclicity. A thickening of the ovarian tunica albuginea and a slight hardening of the ovaries were found postmortem. In conclusion, dairy heifers can occasionally show cyclic activity and form CL-like structures during twice-weekly OPU. Further, OPU did not seem to cause any major negative effects on ovarian structure and subsequent ovarian function.

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Three experimental and three control heifers were included in the study. After a control period (CP), the experimental heifers were subjected to 4 and 5 weeks of twice-weekly follicular punctures (FPP1 and FPP2) with two oestrous cycles in between. The follicular punctures were performed using the ovum pick-up (OPU) technique. During the CP and FPPs, ovarian activity was monitored by transrectal ultrasonography and analyses of plasma progesterone, prostaglandin F2alpha (PGF2alpha) metabolite, luteinizing hormone (LH), oestradiol and follicle stimulating hormone (FSH) levels. The animals were slaughtered 7 days after cessation of FPP2 and their ovaries examined. The control animals were monitored for their oestrous cyclicity and were then ovariectomized and the ovaries examined. Despite the absence of ovulation, the formation of corpus luteum (CL)-like structures could be ultrasonographically observed during FPPs. These structures either had characteristics comparable to those of the CLs during the CP (group 1) or were smaller and had a shorter life span and (or) luteal active phase with a lower progesterone production (group 2). In group 1, CL-like structures emanated from punctures of large-sized follicles on or close to the days on which the animals showed oestrus and had a rise in oestradiol and LH. In group 2, the structures developed from punctures of smaller-sized follicles and no oestrus nor a detectable 1.14 rise was revealed around punctures. During most FPPs, the basal level of LH was at zero. During all FPPs, a significant rise in FSH level occurred on the day following the day of puncture. To conclude, repeated follicular puncture appeared to alter slightly endocrine profiles and cause minor morphological changes in the ovaries. Both the macro- and microscopic examinations revealed the presence of luteal structures and follicles of various sizes in the ovaries of all heifers studied. The connective tissue in the ovarian tunica albuginea of the experimental heifers was significantly thicker than that in controls (P < 0.001).

The aim of the present study was to assess the effects of the ovum pick-up (OPU) technique on animal well-being. Eight dairy heifers were subjected to 4 months of twice-weekly OPU. The physiological response to OPU was recorded in four heifers at two sessions, at the beginning (time 1) and at the end (time 2) of the 4-month period. Heart rates were measured and blood was analysed for cortisol, vasopressin and PG-metabolite before, during (every 5 and 2(1/2) min), and after the OPU sessions. Reactions to each subprocedure of OPU ('restraint', 'epidural', 'device in' and 'puncture') were closely observed. In all heifers, reactions to the OPU procedures were also noted throughout the experimental period, and changes in routine behaviour, oestrous behaviour, body temperature, or other clinical traits were recorded. Subsequent to the experiment, the ovaries and tails were carefully inspected. At time 1, there was an insignificant increase in heart rate and cortisol throughout the OPU procedure. At time 2, these two parameters increased significantly, but both parameters declined to pre-OPU levels 10 min after completion of the procedure. No significant changes were seen in vasopressin or PG-metabolite at time 1 and time 2. Behaviourally, the heifers showed the strongest response to epidural anaesthesia, with a tendency for more intense response during the late 4-month sessions. The response to 'device in' and 'puncture' varied among individuals independently of time. There were no changes in the routine or oestrous behaviour throughout the experiment and no signs of clinical disorders. No major pathological changes were macroscopically seen in the ovaries and tails subsequent to the 4 months of OPU. In conclusion, the heifers showed a response to OPU, mostly to administration of epidural anaesthesia. However, we demonstrated that epidural anaesthesia can be administered in a way causing less discomfort.

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Repeat-breeder heifers (RBH) have been shown to present reproductive perturbations during spontaneous cyclicity, which affects oestrus and ovulation. Some of these disturbances (e.g. deviating hormone patterns) are also present during and after cycles of twice-weekly ovum pick-up (o.p.u.), performed according to an optimized schedule allowing normal oestrous cyclicity. In the present study, the effects of o.p.u. on oocyte competence in in vitro maturation (i.v.m.) and in vitro fertilization (i.v.f.) have been evaluated, as were the effects on expression of oestrus and fertility in five RBH (> or =4 artificial inseminations) and five virgin heifers (VH controls). In total, 269 RBH and 174 VH oocytes were scored for quality prior to i.v.m. and i.v.f. The number of follicles available for puncture was higher in RBH, but the oocyte recovery rate after o.p.u. was lower in RBH compared with VH controls and the recovered RBH oocytes were of lower quality, as judged by their appearance at retrieval. Confocal laser scanning and transmission electron microscopy of immature oocytes did not reveal any differences between RBH and VH control oocytes with respect to nuclear and mitochondrial status. However, after i.v.m., the cytoplasmic spatial reorganization of mitochondria and cortical granules was less advanced in RBH, which could contribute to the subfertility that defines the syndrome. Cleavage rates after i.v.f. were similar in RBH and VH controls. Subsequent to the o.p.u. period, in vivo fertility after controlled artificial insemination was comparable with field fertility rates in both RBH and VH.