Raymond J. Walsh

The distribution of lactogen receptors in the mammalian hypothalamus: an in vitro autoradiographic analysis of the rabbit and rat

The hypothalamus contains a high concentration of lactogen receptors as detected with in vitro radioreceptor assay techniques. In an effort to define the location of the lactogen receptors relative to specific hypothalamic nuclei, an in vitro autoradiography technique was applied to frozen sections of rat and rabbit brains. Three lactogenic hormones, i.e. human growth hormone (hGH), ovine prolactin (oPRL), and rat prolactin (rPRL), were radiolabeled with iodine-125. Competition for observed binding sites was assessed with unlabeled hGH, oPRL, and bovine growth hormone (bGH). Analysis of the autoradiographs with a microcomputer-based densitometry system revealed that the rabbit hypothalamus contains specific lactogen binding sites within the supraoptic, paraventricular, suprachiasmatic, ventromedial, arcuate, and dorsomedial nuclei and the medial preoptic area. Unlabeled bGH was effective in competing for binding sites in all areas when hGH but not oPRL was used as the radiolabeled ligand, suggesting the presence of growth hormone receptors in the rabbit hypothalamus with a distribution similar to that of the lactogen binding sites. In contrast to the rabbit, no lactogen binding sites were detected in the rat hypothalamus regardless of the ligand used in the assay. All of the ligands were successful, however, in detecting lactogen receptors within the rat choroid plexus and liver. The results from the rabbits indicate that the influences of prolactin on hypothalamic activity are mediated via lactogen receptors that are widely distributed throughout the various pertinent hypothalamic nuclei. The broad distribution of lactogen receptors in the rabbit hypothalamus attests to the extensive influence of prolactin on hypothalamic regulatory systems. The results from the rat raise questions as to the nature of rat brain prolactin receptors in comparison to prolactin receptors in rat peripheral tissues.

Virtual reality surgical simulation for lower urinary tract endoscopy and procedures.

BACKGROUND AND PURPOSEnTo provide a realistic experience of lower urinary tract endoscopic procedures, we have developed and continue to expand a computer-based surgical simulator that incorporates a surgical tool interface with anatomic detail and haptic feedback.nnnMETHODSnSurface-based geometric data for the lower urinary tract were generated from the National Library of Medicine Visible Human dataset. The three-dimensional texture map of the surface geometry was developed from recorded endoscopic video procedures. Geometry and associated texture maps were rendered in real time using the Silicon Graphics Extreme Impacts program. The surgical interface device incorporated all normal ranges of motion and resistance that occur within an actual operative environment. The hands-on endoscopic device attached to the interface device was provided by Circon-ACMI, Inc. Urologic residents evaluated the program for correlation with actual endoscopic procedures.nnnRESULTSnTexture-mapped digitized images provided a close anatomic similarity to actual videoendoscopic images. Virtual endoscopy of the lower urinary tract was reproducible and closely simulated actual visual and tactile endoscopic experience.nnnCONCLUSIONSnVirtual reality surgical simulation is feasible for a variety of lower urinary tract procedures. This system coordinates visual perception with appropriate haptic feedback in both longitudinal and rotational axes. These types of procedures may be incorporated into future educational experiences for urologists to introduce new techniques and to provide documentation of surgical experience.

The ontogeny of specific prolactin binding sites in the rat choroid plexus

The development of prolactin receptors in the choroid plexus of the rat was examined using the in vivo autoradiographic approach employing the principle of competitive binding. Experimental animals were injected with [125I]prolactin alone (total binding) while control animals received [125I]prolactin and a 500-fold excess of unlabelled prolactin (non-specific binding). Newborns as well as animals 10, 14 and 18 days postnatal were studied. Three minutes following hormone injection animals received an intracardiac perfusion with fixative and tissues were prepared for quantitative light microscopic autoradiography. The choroid plexus first demonstrated specific binding of prolactin, i.e. a statistically significant difference in the autoradiographic reactions between experimental and control animals, at 14 days postnatal. The lactogen specificity of these binding sites was further defined by the ability of [125I]prolactin to be displaced by unlabelled human growth hormone, which is lactogenic in rats, and not by unlabelled insulin, which is structurally dissimilar to prolactin. Morphometric analyses were performed on electron micrographs of choroid plexus from 10- and 14-day postnatal rats. The volume densities of constituents known to be enriched in polypeptide hormone receptors were measured and compared. Small cytoplasmic vesicles and tubules were statistically significantly more abundant in 10-day-old rats than in 14-day-old animals. It is conjectured that these vesicles and tubules contain an intracellular pool of prolactin receptors whose decrease at 14 days parallels the expression of specific binding sites at the cell surface.

In vivo autoradiographic analysis of prolactin binding in brain and choroid plexus of the domestic ring dove

SummaryThe binding of intravenously administered prolactin to choroid plexus and brain tissue was determined radioautographically in the ring dove, a species that exhibits prolactin-induced alterations in brain function. An intense autoradiographic reaction was detected over the epithelial cells of the choroid plexus 5 min after the intravenous injection of 125I-ovine prolactin. A significant reaction was also observed over the infundibulum but no significant uptake of prolactin occurred in other brain areas. The binding of radiolabelled prolactin to infundibulum appeared to be non-specific, since excess unlabelled hormone did not reduce silver grain density. In contrast, 125I-ovine prolactin binding in choroid plexus was significantly reduced by excess unlabelled ovine prolactin or human growth hormone, but not by ovine luteinizing hormone. Specific binding to choroid plexus was also detected in vitro. The lack of significant brain uptake of prolactin in vivo is discussed in relation to recent in vitro evidence for specific binding sites for prolactin in several dove brain regions. Similarities between the binding results obtained in this avian species and those reported previously in mammals suggest that the two vertebrate groups exhibit similar patterns of prolactin interaction with neural target tissues.

Prolactin receptor localization to the area postrema.

The area postrema, which lacks a blood-brain barrier, was examined for the presence of prolactin receptors, which would render it a potential site for vascular prolactin to directly interact with neuronal elements. Using an in vitro autoradiographic technique, frozen sections of New Zealand white rabbit medulla were incubated with radiolabelled ovine prolactin alone (total binding) or radiolabelled ovine prolactin in the presence of excess unlabelled ovine prolactin (non-specific binding). The specificity of the binding was also assessed using excess unlabelled human prolactin or ovine LH. While excess unlabelled ovine and human prolactin caused a statistically significant reduction in radio labeled prolactin binding, unlabelled LH was without effect. Results reveal the presence of specific prolactin binding sites within the area postrema, a previously unknown prolactin target area of the CNS.

Distribution of substance P positive cells and nerve fibers in the rat thymus

The immune and nervous systems communicate through an array of signalling molecules which includes substance P. This work investigates the anatomical relationship between substance P nerve fibers, receptors, and substance P positive cells in the thymus. Thymuses from rats were frozen or paraformaldehyde fixed. In vitro autoradiography was used to map the distribution of SP receptors. Immunostaining was used to localize SP positive cells and nerve fibers by transmitted light and confocal microscopy. SP receptors exhibited a broader distribution than previously reported, being present throughout the organ with a preferential concentration in the cortico-medullary zone. While SP fibers were frequently associated with the blood vasculature, they were also present throughout the organ independent of blood vessels and were most prominent in the cortico-medullary zone. SP positive cells followed a similar pattern of distribution as the SP fibers and were present as single cells or aggregates of SP positive cells. Confocal microscopy revealed close spatial contact between the SP positive nerve fibers and the SP positive thymic cells. The close spatial relationship between the SP positive thymic cells and SP positive nerve fibers supports the concept of a structural-functional unit between SP nerve fibers and their potential receptor-bearing target cells in the thymus.

Distribution of specific substance P binding sites in the heart and adjacent great vessels of the Wistar white rat

Abstract.Magnesium(Mg)-deficiency, whether dietary or an effect of a clinical condition such as diabetes, results in a variety of cardiovascular pathologies. Substance P (SP) has been implicated in the induction of cardiac focal inflammatory lesions that occur during Mg-deficiency. Blockade of SP receptors results in a significant reduction in the incidence of lesion formation. In an effort to identify potential endogenous cell populations of the heart, which may play a role in SP-dependent lesion formation, film- and light-microscopic autoradiography were used to map the distribution of specific SP binding sites in frozen sections of the normal rat heart and adjacent great vessels. Binding was assessed with 0.1u2005nM I-125 Bolton-Hunter labelled SP in the absence (total binding) or presence (non-specific binding) of excess unlabelled SP, prolactin, or L-703,606, a non-peptide antagonist of SP receptors. Film autoradiograms revealed prominent small foci of intense autoradiographic reactions dispersed intermittently around the periphery of the great vessels and coronary arteries, among the interstitial connective tissue of the heart, and along the cusps of the cardiac valves. Excess unlabelled SP caused a significant reduction (97.7% displacement; P<0.001) in the focal autoradiographic reactions. L-703,606 caused a similar reduction in SP binding (97.3% displacement; P<0.001), while prolactin had no statistically significant effect on the binding of radiolabelled SP. Light-microscopic autoradiograms revealed that the SP binding sites occurred within clusters of connective tissue cells or in rarely observed parasympathetic ganglia. No evidence was found to suggest the presence of SP receptors on endothelial cells, cardiac muscle fibers, or smooth muscle fibers. The connective tissue cells which bound SP within the heart will likely include types that are susceptible to SP activation and thus may play a role in initiation of the focal inflammation characteristic of Mg-deficiency.

Magnesium-dependent non-specific binding of [125I]prolactin to myelinated tracts in the rat central nervous system.

An in vitro autoradiographic assay was used in identifying a magnesium-dependent, non-specific binding of [125I] prolactin to myelinated fiber tracts in the rat brain. Frozen tissue sections were incubated for 18 h at 4 degrees C in media which included [125I]prolactin alone or with a 500 fold excess of unlabelled prolactin. Magnesium in the incubation medium caused a non-specific binding of radiolabelled prolactin to the myelinated fiber tracts in the brain. In contrast, calcium did not facilitate prolactin non-specific binding to myelin. Hence, calcium should optimize the detection of specific prolactin binding sites in the brain by in vitro autoradiographic or radioreceptor assays.

Virtual Reality Surgical Simulation for Lower Urinary Tract Endourologic Surgery

Computer-based simulations for clinical training have been utilized for medical education for over 25 years, with progression of technological advances from mainframe computers to the multimedia-based systems in current use.5 Simulation becomes attractive whenever actual physical experience is associated with unacceptable risk or expense or the experience is not practical because of distance or temporal constraints. The classic example of the value of simulation in such circumstances is flight simulation aviation training, which was first introduced during the Second World War to reduce the high casualty rate among the relatively inexperienced pilots recruited for aerial warfare. Though its development has occurred over 40 years, flight simulation in a virtual environment has only recently evolved sufficiently to be considered indispensable for training, due to the savings and benefits in time, expense, equipment, and safety it offers.

Active illumination based 3D surface reconstruction and registration for image guided medialization laryngoplasty

The medialization laryngoplasty is a surgical procedure to improve the voice function of the patient with vocal fold paresis and paralysis. An image guided system for the medialization laryngoplasty will help the surgeons to accurately place the implant and thus reduce the failure rates of the surgery. One of the fundamental challenges in image guided system is to accurately register the preoperative radiological data to the intraoperative anatomical structure of the patient. In this paper, we present a combined surface and fiducial based registration method to register the preoperative 3D CT data to the intraoperative surface of larynx. To accurately model the exposed surface area, a structured light based stereo vision technique is used for the surface reconstruction. We combined the gray code pattern and multi-line shifting to generate the intraoperative surface of the larynx. To register the point clouds from the intraoperative stage to the preoperative 3D CT data, a shape priori based ICP method is proposed to quickly register the two surfaces. The proposed approach is capable of tracking the fiducial markers and reconstructing the surface of larynx with no damage to the anatomical structure. We used off-the-shelf digital cameras, LCD projector and rapid 3D prototyper to develop our experimental system. The final RMS error in the registration is less than 1mm.