Ann-Judith Silverman

The luteinizing hormone-releasing hormone (LHRH) systems in the rat brain.

Immunocytochemical procedures on thick, unembedded sections were used to visualize the neurons and their processes that contain LHRH-immunoreactive material in the rat central nervous system (CNS). In animals pretreated with colchicine (75 micrograms, intraventricularly), cell bodies could be observed as far anterior as the olfactory bulb and posterior to the retrochiasmatic area of the basal hypothalamus. Several new observations for the rat were made in this study, including LHRH neurons in the accessory olfactory bulb and other olfactory-related structures, and in the anterior hippocampus and the induseum griseum. As in studies from other laboratories, we observed many LHRH cells in the periventricular medial preoptic area, diagonal band of Broca and septal nuclei, and fewer positive cells in the anterior hypothalamic area and the region of the supraoptic commissure. The LHRH fibers from all of these cells are widely dispersed in the CNS. In addition to the dense innervation of the median eminence, positive fibers are found innervating other circumventricular organs, coursing close to the ependymal wall of the ventricular system or in close association with cerebral arteries and areas of the pia mater and subarachnoid space. LHRH fibers may also innervate neurons in several regions of the CNS. A novel projection of LHRH fibers for the rat was found originating from supracallosal neurons and coursing through both cingulate and neocortex. The possible distribution of efferents from each LHRH cell group is discussed.

Comparative Distribution of Vasopressin and Oxytocin Neurons in the Rat Brain Using a Double-Label Procedure

The distribution of vasopressin (VP) and oxytocin (OT) neurons in the rat supraoptic (SON), paraventricular (PVN), and accessory magnocellular (AMN) nuclei was studied by localizing both peptides on the same section with a double immunocytochemical staining procedure employing specific monoclonal antibodies (MAB). This procedure allows us to visualize the distribution of one cell type relative to the other. In the rostral SON, VP cells lie dorsal and medial to the OT cells. Near the mid-point of the nucleus along its rostral-caudal length, there is a transition zone in which the two cell types are mixed. Proceeding caudalward, the relative locations of OT and VP cells are exchanged so that most of VP cells are located in the ventral and medial sector of the nucleus, whereas the OT cells are situated dorsal and lateral. However, there is no absolute segregation of the two types of cells anywhere in the nucleus. In the anterior part of the PVN a rostral group (rPVN) of cells composed of a medial portion and a lateral wing can be recognized. Nearly all of the cells in the rPVN are oxytocin-containing. The rPVN is separated from the next group, the middle PVN (mPVN), by a cell poor zone of about 100-150 micron. The mPVN contains both OT and VP neurons. As one proceeds caudally, the OT cells extend in the rostrocaudal direction from an anterior and ventromedial location, forming a shell around a core of VP neurons. In the most caudal PVN (cPVN), a triangular cell group characterized by fusiform cells with long-beaded processes can be distinguished from the more rounded cells of the remaining PVN. Many fusiform cells in the cPVN appear to send their axons to the posterior perifornical nucleus and the nucleus of the medial forebrain bundle. Other fusiform cells of the cPVN are oriented in a rostral-caudal plane and are situated more medially in this subdivision. The dendrites of these cells project into the mPVN while their posterior processes, most of which also appear to be dendrites, project caudally along a medial route.

Light and Electron Microscopic Immunocytochemical Analysis of Antibodies Directed Against GnRH and Its Precursor in Hypothalamic Neurons

A battery of antibodies directed against different portions of the precursor to gonadotropin-releasing hormone (GnRH), as well as to the mature decapeptide, were characterized immunocytochemically in two ways. Absorption experiments were used to determine the epitope recognized by each antiserum. Electron microscopic immunocytochemistry was then used to define the subcellular organelles that contained reaction product when tissue was incubated with these reagents. These latter observations helped to determine if the antibody recognized the epitope as part of the intact precursor or only after it had been cleaved from parent protein. Our results demonstrate that the GnRH precursor is routed from the rough endoplasmic reticulum through the Golgi apparatus to the secretory vesicles. Furthermore, we show that initial cleavage and processing of the GnRH precursor begin in the cell soma. These antibodies should be useful in the future in determining changes in processing of precursor in animals that differ in endocrine function.

Modification of Hypothalamic Neurons by Behavioral Stress

The pituitary-adrenocortical and sympathetic-adrenomedullary systems, under neural control of select cell populations in the hypothalamus, coordinate the broad profile of adaptive bodily responses that collectively define the emergency reaction of the organism. To meet the threat of an environmental stressor, the body is initially readied for action: heart rate, blood pressure and respiration are increased, muscles function more efficiently, pain sensitivity is dampened, and a variety of other responses are coordinated in what Selye1 termed the alarm reaction. Once the threat is reduced or identified (for all such responses display adaptation to repeated exposures to the same brief stressor), the action of these systems is self-limiting, and the body returns to normal. However, some stress situations are chronic, or recur periodically in a pattern that defeats adaptation. As a consequence of this type of malignant exposure to stress, certain bodily functions fail to return to pre-stress levels and remain in a prolonged activated state, overriding endogenous homeostatic mechanisms.

Hormonal effects on development of transplanted embryonic hypothalamus

Abstract Transplantation of the embryonic hypothalamus onto the choroidal pia of an adult female rat host offers unique opportunities for studies of the humoral and neural factors which regulate hypothalamic differentiation and the acquisition of neuroendocrine function. Our previous work established that the transplanted hypothalamus acquires immunoreactivity for a number of peptides (e.g., LHRH, somatostatin, neurophysin), the ability to sequestor 3 H-estradiol, and an organized median eminence-like structure with catecholamine fluorescence. This report presents initial data on three additional aspects of the hypothalamic transplants which are important to assess their ability to function: (1) Estrogen receptors are characterized and measured biochemically in transplants and their elaboration following transplantation is shown to be independent of the presence of gonadal steroids through the use of gonadectomized hosts. Estrogen-mediated induction of progestin receptor sites, a feature of the normal mature hypothalamus, is shown to occur in hypothalamic transplants in both intact and gonadectomized hosts. (2) Neurophysin immunostaining also develops in a seemingly normal manner in intact and in gonadectomized and adrenalectomized (GDX-ADX) hosts given corticosterone. However, preliminary observations on transplants into GDX-ADX hosts without corticosterone replacement suggest that glucocorticoids, which are present in the maternal and fetal blood, may increase the number and/or immunoreactivity of neurophysin-containing perikarya. (3) Initial experiments on bone growth in hypophysectomized hosts indicate that pituitary-hypothalamic cotransplants function to some extent to promote bone growth above the low level seen after total hypophysectomy of the host. It is unknown at this time the degree to which the hypothalamic cotransplant functions to drive the pituitary transplant.

Neuroendocrine brain grafts

Neuroendocrine grafts have been used to ameliorate symptoms associated with genetic or imposed deficiencies. In the course of these studies, it has become evident that the technique may be used to study some basic questions in neurobiology, including targeting, plasticity and integration of neurons. Among the genetic models employed, the hypogonadal mouse, deficient in gonadotropin-releasing hormone (GnRH), has provided an excellent opportunity to address certain of these issues. GnRH cells present in preoptic area grafts derived from normal mice show directed axonal outgrowth to the median eminence of the host, and GnRH neurons may migrate from the graft into the host hypothalamus. The adult host brain sends identified neuronal processes, into the graft and modulates physiological functioning of the grafted cells.

Functional GnRH Neuronal Transplants in the Hypogonadal Mouse

Transplantation of neuronal tissue containing GnRH cells into the brain of the mutant hypogonadal mouse has provided us with an important tool to study mechanisms of GnRH secretion. Lacking GnRH because of a deletion in the GnRH gene (1), the adult hpg mouse has an undeveloped reproductive system (2), but responds to implantation of normal fetal or neonatal preoptic area tissue grafts into the third ventricle of the brain with reproductive development. GnRH cells survive within the grafts (Fig. 9.1a) and innervate the median eminence of the host brains (Fig. 9.1b). Both male (3, 4) and female (5) hpg mice with grafts (hpg/POA) increase pituitary gonadotropin production, which results in gonadal development.