Naomi E. Rance, MD University of Maryland 1983, PhD, University of Maryland 1981 - US grants

Our overall goal is to characterize and understand the events that occur in the human central nervous system in response to menopause. We have found that human menopause is characterized by a striking hypertrophy of neurons within the hypothalamic infundibular nucleus. Postmenopausal neuronal hypertrophy occurs in a subpopulation of neurons containing estrogen receptor and neurokinin B (NKB) mRNA and is accompanied by a marked increase in tachykinin gene expression. GnRH gene expression also increases, but in a separate subpopulation of hypothalamic neurons. During our last funding period, we used animal models to provide strong evidence that the changes in NKB cell size and gene expression in postmenopausal women are due to removal of steroid negative feedback. In this renewal, we propose the first studies to elucidate the physiological relevance of the increase in NKB gene expression in postmenopausal women. In specific aim 1, we will test the hypothesis that NKB neurons participate in the hypothalamic circuitry regulating LH secretion in the rat using pharmacological tools and antisense oligodeoxynucleotide (ODN) knockdown techniques. In specific aim 2, we will characterize the anatomic relationship between arcuate NKB neurons and the reproductive axis. We will determine if arcuate NKB or NK3 receptor-immunoreactive neurons project to the preoptic-septal region (site of GnRH neurons) or the primary capillary plexus of the median eminence in the rat. We will also determine if GnRH neurons express NK3 receptor-immunoreactivity in the human or rat hypothalamus. In specific aim 3, we will attempt to gain a deeper understanding of the remarkable changes in the infundibular nucleus of postmenopausal women. We will determine if menopause is association with signs of cellular degeneration such as increased microglial cells or GFAP gene expression. We will also use the Golgi-method to determine if the neuronal hypertrophy is associated with expansion or regression of infundibular dendritic arbors. Our studies provide an exceptional opportunity to study human brain specimens and address basic biological, issues directly relevant to the physiology of postmenopausal women.

DESCRIPTION (provided by applicant): One of the most prominent symptoms of menopause is the hot flush, a disorder of central hypothalamic thermoregulation. Despite the unprecedented numbers of individuals affected, there is little understanding of the etiology of flushes and few laboratories are studying the basic biology of this phenomenon. Hot flushes are caused by estrogen withdrawal and consist of the coordinated activation of the physiologic mechanisms to dissipate heat (cutaneous vasodilatation, sweating and behavioral changes). Integration of the hypothalamic control centers for thermoregulation and reproduction is likely to be a key element in the mechanism of flushes. In our previous studies, we described dramatic changes in the morphology and gene expression of estrogen receptor-containing NKB mRNA containing neurons in the infundibular (arcuate) nucleus of postmenopausal women. Extensive evidence gathered in animal models has shown that the increase in NKB gene expression in postmenopausal women is secondary to loss of ovarian estrogen. We hypothesize that these NKB neurons play a role in the activation of thermoregulatory vasodilatation and therefore could be involved in the generation of menopausal flushes. We have now collected substantial pilot data in support of this hypothesis and have evidence of a novel neural circuit whereby estrogen-withdrawal activates heat dissipation effectors. We postulate that estrogen-responsive NKB neurons in the rat arcuate nucleus activate thermoregulatory vasodilatation via projections to NK3 receptor- expressing neurons in the median preoptic nucleus (MnPO), an important regulatory center for autonomic function. The present proposal will focus on this hypothesis. In specific aim 1, we will use morphologic tools to examine relationship between estrogen-responsive NKB neurons in the rat arcuate nucleus and the CNS centers controlling thermoregulatory vasodilatation. Specific aim 2 will test the hypothesis that NK3 receptor activation of MnPO neurons stimulates heat dissipation effectors in the rat. Specific aim 3 will evaluate whether interference with the function of NK3 receptor-expressing MnPO neurons reduces tail skin temperature or alters the thermoregulatory axis in ovariectomized rats. Specific aim 4 will determine if NK3 receptor mRNA is expressed in neurons in the human MnPO. These studies will provide novel insights into the potential relationship between the changes in hypothalamic gene expression in postmenopausal women and the inappropriate activation of heat dissipation mechanisms that characterize the hot flush. Understanding the mechanisms of hot flushes is critical for the ultimate design of appropriate therapies. A major feature of menopause is the hot flush, which is considered to be a disorder of central nervous system thermoregulation. Hot flushes are caused by loss of estrogen and are characterized by inappropriate activation of the physiological systems that remove heat from the body (heat dissipation). Currently, no safe and efficacious treatment is available, despite the fact that hot flushes may severely impact the quality of life in many individuals. In our previous research studies, we discovered that a group of neurons expressing the peptide neurokinin B (NKB) exhibit increased gene expression in the hypothalamus of postmenopausal women. We hypothesize that these NKB neurons play a role in the physiologic mechanisms to dissipate heat and therefore could be involved in the generation of hot flushes. The present proposal is designed to test this hypothesis. PUBLIC HEALTH RELEVANCE: These studies would be the first to relate the changes in hypothalamic gene expression in postmenopausal women to one of the most prominent symptoms of menopause. Little is understood about the mechanisms of hot flushes and there are only a few laboratories that study the basic biology of this problem. Understanding the mechanisms of hot flushes would greatly facilitate the design of appropriate treatments for these symptoms.

DESCRIPTION (provided by applicant): Hot flushes occur in the majority of menopausal women and in young women after estrogen withdrawal. They are characterized by the activation of heat dissipation effectors, including skin vasodilatation, sweating, and cold-seeking behavior. Despite the millions of individuals who experience these symptoms, the cause remains an enigma, and there is little understanding of the neural circuits for estrogen modulation of body temperature. Our laboratory has long hypothesized that kisspeptin/neurokinin B/dynorphin (KNDy) neurons contribute to the generation of flushes, because of their dramatic changes in the hypothalamus of postmenopausal women. In support of this hypothesis, we recently showed that ablation of KNDy neurons in the rat decreases cutaneous vasodilatation and alters the effects of estrogen on thermoregulation. The goal of the present grant is to elucidate the downstream (preoptic) pathways used by estrogen-responsive KNDy neurons to mediate thermoregulatory vasodilatation. KNDy neurons project to key thermoregulatory structures that regulate heat- dissipation effectors: the median preoptic nucleus (MnPO) and medial preoptic area (MPO). Moreover, both these areas express the primary NKB receptor (NK3R) and activation of NK3R in the MnPO reduces body temperature. We hypothesize that NK3R neurons in the MnPO and MPO integrate information from estrogen-responsive KNDy neurons with warm thermal- signals that trigger heat dissipation effectors. The following specific aims are proposed: 1) To evaluate whether NK3R neurons in the MnPO and MPO of Tacr3-EGFP mice are activated by warm-thermal signals from the environment, are modulated by estrogen and receive inputs from KNDy neurons; 2) To perform electrophysiological recordings of NK3R neurons in the preoptic area of the Tacr3-EGFP mouse to determine if they are warm-sensitive, receive inputs from KNDy neurons and if their thermal sensitivity is altered by estrogen or NK3R signaling; 3) To determine if NK3R neurons in the MnPO and MPO are essential for the estrogen modulation of body temperature; 4) To determine if a homologous projection pathway from KNDy neurons to preoptic NK3R neurons exists in the human. These studies will shed light on the integration of reproductive and thermoregulatory systems and provide clues into the etiology of hot flushes. Understanding the mechanisms of hot flushes is essential for designing targeted therapies.