No abstract

Parathyroid tumors occur either sporadically or as part of inherited syndromes such as multiple endocrine neoplasia (MEN) types 2A and 2B. The development of both of these familial syndromes has been related to specific germline gain-of-function mutations predominantly in exons 10 and 11 (MEN 2A) and 16 (MEN 2B) of the RET proto-oncogene. The same mutations have also been implicated in the pathogenesis of sporadic medullary thyroid carcinoma and sporadic pheochromocytoma. The RET mutations are thought to have a transforming effect only in cells of neural crest origin such as thyroid parafollicular (C-cells) and adrenal chromaffin cells, which normally express the RET proto-oncogene. Expression of RET messenger RNA has not yet been studied in the parathyroid, however, we demonstrate in this study by a sensitive, semiquantitative RT-PCR technique and in situ hybridization, that RET is expressed in MEN 2A parathyroid tumors and in sporadic adenomas. Although DNA from a parathyroid tumor of a MEN 2A patient displayed an expected mutation, none of the previously described MEN 2A or 2B mutations were found in DNA of 34 sporadic adenomas. Our data suggest that parathyroid disease is an integral part of the MEN 2A syndrome, but that MEN 2 mutations in RET rarely play a part in the pathogenesis of sporadic parathyroid tumors.

In this study we investigated the presence and anatomical location of atrial natriuretic factor (ANF) receptor subtypes in the rat central nervous system using in vitro autoradiographic and cross-linking techniques.

In vitro autoradiographic studies showed that high-affinity atrial natriuretic factor (ANF) binding sites are present on rat choroid plexus (Kd = 83.8 pM, B_max = 22.9 fmol/mg protein). Guanylate cyclase-coupled receptors (ANF-Ri) represent 30% and non-guanylate cyclase-coupled ANF receptors (ANF-R2) represent 70% of the total ANF receptors present in this tissue. To provide detailed cellular localization of the binding sites, the technique of electron-microscopic autoradiography was applied using ^l25I-ANF (Ser 99-Tyr 126) as an in vivo ligand. In order to identify possible binding sites at the basolateral and the apical aspects of the choroid plexus, the ligand was injected into the carotid artery or into the lateral cerebral ventricles, respectively. Light-microscopic autoradiography demonstrated that ANF binds specifically to choroid plexus regardless of its route of administration. Electron-microscopic autoradiography showed that silver grains were localized primarily on epithelial cells of the choroid plexus (96–99%) and marginally on endothelial and pial cells. In choroidal epithelial cells, ultrastructural analysis of silver grain distribution revealed that, at 2 min after intracarotid or intracerebroventricular ¹²⁵I-ANF injection, lysosomes were the most distinctly labeled organelle (highest relative specific radioactivity). HPLC Chromatographic analysis disclosed that 96–99% of choroid plexus-bound ANF was already degraded 2 min after injection and that at least 63–66% of the degradation took place at the plasma membrane. These results indicate that ANF binding sites are present on both aspects of choroidal epithelium, and suggest that ANF is very quickly degraded in choroid plexus by membrane-associated as well as lysosome-associated processes.

The area postrema (AP) is a brain stem circumventricular organ implicated, among other functions, in central cardiovascular (CV) regulation. Competition binding analysis performed by quantitative in vitro autoradiography demonstrated specific, high-affinity (Kd, 0.32 +/- 0.11 nM), low-capacity (Bmax, 57.5 +/- 10.9 fmol/mg protein) atrial natriuretic factor (ANF) binding sites in the AP. C-ANF [des-(Gln116-Gly120)ANF-(Arg102-Cys121)-NH2] and ANF-(Phe106-Ile113)-NH2 (two ligands endowed with selectivity for the ANF-C receptor), as well as C-type natriuretic peptide (CNP), did not compete noticeably at pathophysiological concentrations for 125I-ANF binding. 125I-[Tyr0]CNP bound to the AP to a much lower extent than 125I-ANF. Electron microscopic autoradiography in vivo disclosed that 125I-ANF was preferentially bound to axon, dendrite, and astrocyte plasmalemma. These studies demonstrate that the AP contains natriuretic peptide binding sites with pharmacological characteristics of the ANF-A and ANF-B but not of the ANF-C receptor subtype. In the AP, ANF interacts with those sites resembling ANF-A receptors. Cellular localization of these binding sites may relate to their possible involvement in the centrally mediated salt and water regulation and/or CV effects of circulating ANF.