Krajewski-Hall SJ, Mittelman-Smith, Mcmullen NT, Rance NE (2013) Ablation of arcuate KNDy neurons amplifies the LH surge in steroid-primed, ovariectomized rats. Neuroscience 2013 Abstracts 274.01. Society for Neuroscience, San Diego, CA.
Summary: KNDy (kisspeptin, neurokinin B and dynorphin) neurons in the arcuate nucleus play an important role in the reproductive axis. We have developed a method to selectively ablate KNDy neurons in the rat using NK3-SAP, a neurokinin 3 receptor agonist conjugated to saporin (Mittelman-Smith, Endocrinology 2012). Ablation of KNDy neurons results in cessation of estrous cycles, ovarian atrophy, a decrease in tonic LH secretion and loss of the rise of serum LH after ovariectomy. Given these profound effects, we tested if we could induce an LH surge in KNDy-ablated rats using a well-established steroid replacement regimen. Rats were maintained on a 14:10 light cycle (lights on at 0500). Using stereotactic surgery, NK3-SAP or Blank-SAP was injected in the arcuate nucleus and rats were allowed to recover for one month before ovariectomy. Seven days after ovariectomy they were implanted with silastic capsules containing 17β-estradiol. Two days later, they were implanted with progesterone capsules (~0830h). Rats were sacrificed in the afternoon at a time previously shown to exhibit peak LH surge levels (~1600h) and the brains were processed for immunohistochemistry. Ablation of KNDy neurons was verified by near complete loss of NKB-immunoreactive neurons in the arcuate nucleus in NK3-SAP rats. At 0830h, tonic levels of serum LH were significantly lower in KNDy ablated rats, consistent with our previous studies. Unexpectedly, the surge in serum LH at 1600h was more than 3-fold higher in NK3-SAP-treated rats compared to Blank-SAP controls (53.5 + 16.5 ng/ml vs 16.5 + 2.1 ng/ml, respectively). To determine if this change was associated with increased activation of GnRH neurons, dual-labeled GnRH-fos immunocytochemistry was performed in rostral hypothalamic sections. There was no significant difference in the total number of GnRH cells counted in 4 matched sections (NK3-SAP, 85.8 + 9.8 vs Blank-SAP, 84.9 + 4.6) or in the percentage of GnRH cells that were activated, as measured by GnRH-fos coexpression (NK3-SAP, 22.4 + 1.8% vs Blank-SAP, 16.6 + 4.9%). In addition, there was no difference between NK3-SAP and Blank-SAP controls in the number of fos-ir cells counted in the AVPV. These data indicate that arcuate KNDy neurons are not required for the induction of an LH surge. The marked increase in the LH surge in KNDy-ablated rats, however, suggests that KNDy neurons are important for regulating the magnitude of the surge.
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