140. Bos Indicus Heifers Have a Reduced Number of Kisspeptin- and Dynorphin-immunopositive Cells in the Arcuate Nucleus Compared to Bos Taurus Heifers.

Abstract Bos indicus-influenced cattle represent approximately 30% of the U.S. herd. While better adapted to tropical and subtropical climates, Bos indicus-influenced females exhibit reduced reproductive efficiency and delayed puberty compared to Bos taurus. These reproductive differences revolve primarily on the neuroendocrine system, however the specific neuronal networks and peptides involved remain unclear. Arcuate nucleus (ARC) KNDy neurons, which co-express kisspeptin, neurokinin B, and dynorphin, regulate GnRH pulsatile secretion and mediate steroid feedback mechanisms. In ruminants, this neuronal population is thought to function as the core of the hypothalamic GnRH pulse generator, controlling episodic LH secretion and ovarian function. Therefore, variation in KNDy neuron abundance or peptide expression may contribute to breed-associated differences in reproductive physiology. We hypothesized that Brahman and crossbred (Bos indicus) heifers have reduced kisspeptin and increased dynorphin immunoreactivity compared to Hereford (Bos taurus) heifers. A total of 21 postpubertal heifers were evaluated: Hereford (n = 6), Brahman (n = 8), and crossbred Brahman × Hereford (F1; n = 7). Heifers were ovariectomized during the follicular phase of the estrous cycle to eliminate the effects of endogenous ovarian steroid fluctuations and subsequently implanted with an estradiol-17β subcutaneous implant to maintain basal circulating estradiol concentrations (3.1 ± 0.6 pg/mL). Five days prior to euthanasia, heifers received two intravaginal progesterone devices (CIDR; Zoetis) to produce luteal levels of circulating progesterone. Hypothalami were collected and processed for dual-label immunofluorescence to evaluate kisspeptin and dynorphin colocalization within the ARC. Brain tissue was sectioned at 50 µm, and two rostral, three middle, and two caudal ARC sections per heifer were analyzed. Kisspeptin-positive, dynorphin-positive, and co-expressing neurons were quantified. Data were analyzed using a one-way ANOVA in JMP (SAS), and differences were evaluated using Tukey’s post hoc test. Overall, 64.4% of kisspeptin neurons co-expressed dynorphin, and 83.5% of dynorphin neurons co-expressed kisspeptin, with no breed effect. However, Brahman heifers exhibited a greater percentage of dynorphin neurons colocalizing kisspeptin in the rostral ARC than Hereford heifers (P = 0.04). The total number of kisspeptin-immunopositive neurons in the middle ARC was reduced (P < 0.05) in Brahman compared to F1 and Hereford heifers, while in the caudal and total ARC, Brahman also had a reduced (P < 0.05) number of kisspeptin-immunoreactive cells than F1 but not differing from Hereford heifers. The total number of dynorphin-immunopositive neurons in the middle and total ARC was reduced in Brahman compared to F1 and Hereford heifers. In conclusion, Bos indicus heifers exhibited fewer kisspeptin- and dynorphin-immunopositive neurons in the ARC compared to Bos taurus or F1 counterparts. These changes in the KNDy neuron system are likely to impact GnRH pulse secretion and contribute to the delayed puberty and altered neuroendocrine sensitivity to gonadal steroids observed in Bos indicus cattle.