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The bulk of the secretion of the subcommissural organ is formed by glycoproteins that appear to be derived from two precursor forms of 540 and 320 kDa. Upon release into the ventricle, these glycoproteins aggregate to form Reissner's fiber. We report the isolation of three cDNA clones from a cDNA library prepared from bovine subcommissural organ RNA, by using an anti-Reissner's fiber serum for immunoscreening. Inserts of 0.7, 1.2, and 2.5 kb were amplified by the polymerase chain reaction, subcloned into pUC18 vector, and sequenced. Although restriction mapping of the three inserts initially suggested that all of them were derived from the same mRNA, sequence analysis showed that a short non-homologous region was present in the 0.7-kb insert when compared with the 1. 2-kb and 2.5-kb inserts, suggesting that they corresponded to two different, although highly homologous, mRNAs. Northern analyses showed a single mRNA species of approximately 9.5 kb present in the subcommissural organ and missing in the choroid plexus, brain cortex, and liver. In situ hybridization confirmed that the expression of the RNA was restricted to cells of the bovine subcommissural organ. Polyclonal antibodies raised against a synthetic peptide, whose amino-acid sequence was deduced from the 2.5-kb cDNA, reacted specifically with the bovine and rat subcommissural organ-Reissner's fiber complex. In immunoblots of bovine subcommissural organ, this antibody revealed the precursor 540-kDa form and its putative processed form of 450 kDa. It is concluded that the cloned cDNA encodes for the major constitutive glycoprotein of Reissner's fiber, here designated as RF-Gly I. The sequenced region of RF-Gly I displays a high degree of homology with some regions of the von Willebrand factor and certain mucins; it also displays two motifs homologous with repeats present in proteins of the spondin family and other proteins. A core sequence of the RF-Gly I repeats suggests that this molecule displays protein-binding properties.

The peroxidase-antiperoxidase immunocytochemical procedure was used to study the distribution of ovine corticotropin-releasing factor (CRF) and arginine vasotocin (AVT) immunoreactivities sequentially in the same sections or in adjacent sections of the brain and pituitary of Catostomus commersoni. It was found that all CRF-immunoreactive (IR) neurons in the nucleus preopticus (NPO) also contained AVT immunoreactivity. Co-localization of both immunoreactivities was also observed in fibres forming the preoptic-pituitary tract and in the neurohypophyseal digitations, the IR-CRF and IR-AVT fibres projecting mainly to the neurointermediate lobe (NIL) of the pituitary. An additional population of exclusively IR-AVT neurons and fibres in the NPO, preoptic-pituitary tract and NIL was also observed. Exclusive CRF-immunostaining was found in neurons of the nucleus lateralis tuberis (NLT), in fibres distributed in some diencephalic nuclei and in the neurohypophyseal digitations in the region of the rostral pars distalis (RPD). These results suggest that CRF- and AVT-like substances, present in NIL fibres (probably originating in the NPO), may have an integrated role in the release of the cell products from the pars intermedia, and that the control of corticotrops in the rostral pars distalis, innervated exclusively by IR-CRF fibres (probably originating in the NLT), does not require a simultaneous presence of CRF- and AVT-like substances.

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Background: The subcommissural organ (SCO) is a highly conserved brain gland present throughout the vertebrate phylum; it secretes glycoproteins into the cerebrospinal fluid (CSF), where they aggregate to form Reissner's fiber (RF). SCO-spondin is the major constituent protein of RF. Evidence exists that the SCO also secretes proteins that remain soluble in the CSF. The aims of the present investigation were: (i) to identify and partially characterize the SCO-secretory compounds present in the SCO gland itself and in the RF of the Sprague-Dawley rat and nonhydrocephalic hyh mouse, and in the CSF of rat; (ii) to make a comparative analysis of the proteins present in these three compartments; (iii) to identify the proteins secreted by the SCO into the CSF at different developmental periods.

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The subcommissural organ of vertebrates secretes glycoproteins into the third ventricle that condense to form Reissner's fiber (RF). Antibodies raised against the bovine RF-glycoproteins reacted with the floor plate (FP) cells of two teleost (Oncorhynchus kisutch, Sparus aurata) and two amphibian (Xenopus laevis, Batrachyla taeniata) species. At the ultrastructural level, the immunoreactivity was confined to secretory granules, mainly concentrated at the apical cell pole. In the rostro-caudal axis, a clear zonation of the FP was distinguished, with the hindbrain FP being the most, or the only (Batrachyla taeniata), immunoreactive region of the FP. In all the species studied, the caudal FP lacked immunoreactivity. Both the chemical nature of the immunoreactive material and the rostro-caudal zonation of the FP appear to be conservative features. Evidence was obtained that the FP secretes into the cerebrospinal fluid a material chemically related to the RF-glycoproteins secreted by the subcommissural organ. Thus, in addition to being the source of contact-mediated and diffusible signals, the FP might also secrete compounds into the cerebrospinal fluid that may act on distant targets.