Appearance of a brain-specific antigen (S-100 protein) in the developing rat brain

Abstract— Thelevel of the S-100 protein, a brain-specific antigen, wasdetermined by quantitative complement fixation in the brain stem and cerebrum of the rat during postnatal maturation. The content was minimal at birth in the brain stem and rose to its adult value by day 25. Although S-100 protein could not be detected in the cerebrum of the 2-day-old rat, adult values were also present by the 25th day of age. Neither single dose X-irradiation with 750 rd to the head at 2 days of age or single dose X-irradiation at 11 days of age affected the adult level of S-100 protein in the brain stem or cerebrum. Similarly, hypophysectomy at 20 days of age had no effect on the subsequent levels of S-100 protein.     

Differential localization of "brain-specific" S-100 and its subunits in rat salivary glands

In the rat, the S-100 antigens in the submandibular gland were found to be immunochemically identical with those in the brain (glial cells) when compared using crossed immunoelectrophoresis. Specific antibodies against the S-100a non-beta and against the S-100 beta subunit were prepared from antibodies against crude S-100 protein and from S-100 components (S-100a and b) by affinity chromatography. In the rat salivary glands a differential distribution of subunit immunoreactivity was clearly evidenced using indirect immunofluorescence. Certain intercalated duct cells of the submandibular gland as well as Schwann cells contained the S-100 beta subunit immunoreactivity exclusively, while other duct cells in parotid, submandibular, and sublingual glands contained S-100a non-beta subunit immunoreactivity. Both subunits were present in astrocytes and ependymal cells. The immunocytochemical localization of alpha and beta subunits is a promising technique for the classification of various types of S-100-containing cells.     

Chronic lead intoxication causes a brain-specific nuclear protein to accumulate in the nuclei of cells lining kidney tubules

A characteristic feature of chronic lead intoxication is the induction of intranuclear inclusion bodies in cells lining kidney proximal tubules. These are relatively insoluble lead- and protein-rich structures which may serve a protective function by sequestering lead. The most abundant protein of isolated inclusion bodies, p32/6.3, has been partially characterized by use of a monoclonal antibody. As predicted by biochemical analysis, p32/6.3 occurs in kidney only in conjunction with lead intoxication and inclusion body formation. It does not accumulate in other tissues as a result of lead exposure. Unexpectedly, p32/6.3 was found to be a constitutive protein of adult brain, occurring primarily in the cerebral cortex. Within this tissue, both neurons and astrocytes contained p32/6.3. The brain p32/6.3 was concentrated in the insoluble nuclear protein or matrix fraction, a localization reminiscent of the intranuclear inclusion bodies from lead-exposed kidney. Brain p32/6.3 was detected in rat, mouse, dog, man, and chicken. In rat brain, the appearance of p32/6.3 was developmentally regulated. Only traces were detected 3 days after birth but within 1-2 weeks adult levels were achieved. The presence in brain of a protein which is involved in a potentially protective response to lead suggests that the brain may also have such a protective mechanism.     

S-100 protein immunoreactivity in mammalian testis and epididymis

The present study deals with immunohistochemical localization of S-100 protein in mouse, bank vole and pine vole testis and epididymis. S-100 protein immunoreactivity was observed in the endothelia of capillaries and lymphatic sinusoids of pine vole testis. A reaction to S-100 protein of the same intensity as that noted in the endothelia of testicular capillaries was found in myoid cells of pine vole and bank vole seminiferous tubules. Moreover, a positive reaction to S-100 protein was observed in bank vole and mouse Leydig cells. In the epididymis, a weaker reaction to S-100 occurred in smooth muscles of pine vole and mouse epididymal duct. Despite difficult interpretation of physiological role of S-100 protein we suggest that it may be a part of the blood-testis barrier. It may also participate in the processes of transcytosis and contractility; its cellular expression is regulated by local factors. However, location of S-100 is not specific to the representatives of the same order.     

S-100-like immunoreactivity in a planarian

Summary The presence of an S-100-like immunoreactivity was investigated in the planarian Dugesia gonocephala. By microcomplement fixation assay, measurable amounts of S-100-like immunoreactive material (0.11g/mg soluble protein) were detected in planarian high-speed supernatants. The index of immunological dissimilarity between ox S-100 and planarian S-100-like immunoreactive material was higher than that previously calculated between ox S-100 and all the vertebrates tested. By the immunohistochemical PAP method, S-100-like immunoreactivity was only detectable in the cilia of the epidermal cells. Although the biological meaning of S-100-like immunoreactivity in planarian remains to be clarified, the present data introduce new perspectives into the investigation of S-100.     

A kallikrein-specific inhibitor in rat kidney tubules.

A kallikrein inhibitor was found in tubules of the rat kidney and purified by chromatography on Sephadex G-100. The molecular weight of the inhibitor, estimated by gel filtration and dodecylsulfate electrophoresis, is about 4700. It inhibits the following kallikreins: porcine submanidbular and pancreatic kallikrein, rat kidney and urine kallikrein, and human urine and plasma kallikrein. An inhibition of bovine trypsin was not observed.     

Electroencephalographic effects of the intracentral administration of antibodies to brain-specific antigen S-100]

The EEG method with the assessment of the rhythm power on analogue computers was used to examine the effects of intracentral administration of antibodies to the brain-specific antigen S-100 (S-100 protein). Intracerebral injections of antibodies to S-100 protein caused an increase of the EEG basic rhythm power in the hippocampus, caudate nucleus and the mesenphalic reticular formation, with the subsequent development of epileptiform activity in these structures.     

Peripheral distribution of nervous system-specific S-100 protein in rat

S-100 protein, a nervous system-specific protein, was determined in a soluble extract of various rat tissues with a sensitive enzyme immunoassay method, which consisted of a solid-phase with immobilized anti-S-100 antibody and the antibody labeled with beta-D-galactosidase from Escherichia coli. The minimum detectable amount of S-100 protein was 3 pg/assay. Central nervous tissues (cerebrum, cerebellum, and brain stem) contained 1.4 to 2.8 micrograms S-100 protein/mg protein, whereas most of the peripheral tissues contained less than 0.05 microgram/ml of the specific protein. However, the level of S-100 protein was high in adipose tissue (0.5--1.1 micrograms/mg) and in trachea (about 0.5 microgram/mg), which involves cartilage. The S-100 protein levels in several tissues were significantly higher in female rats than in males at ages of 5 to 6 weeks.     

A new preparation of S-100 protein from rat and bovine brains

The S-100 nervous system protein was purified from bovine and rat brains by a modification of the original procedure. The main modification consisted in substituting a step of calciumdependent binding of S-100 to a phenyl-Sepharose column for the original step of chromatography on G-200 Sephadex. The proteins were pure as determined by SDS gel electrophoresis. HPLC on a reversed phase and on a size-separation column, and by immunological criteria. The bovine S-100 behaved as previously described, during calcium binding, by displaying a conformational change as evidenced by increase in native fluorescence.     

S-100 protein in the testis

Summary S-100, a protein originally believed to be unique to the nervous system, has recently been found in extraneural cell types. We report here on the presence of S-100 in the testis, namely in Leydig cells and in lymphatic endothelial cells, using immunohistochemical and immunochemical methods. We show that the protein in the testis is immunologically identical to brain S-100. The S-100-labelled cells in the testis exhibit morphological similarities with other cell types in different tissues known to contain S-100.     

Immunocytochemical localization of S-100 protein in the brain of adult rat

Summary The cellular and subcellular distribution of the nervous system-specific S-100 protein has been investigated in the brain of adult rat at the ultrastructural level by the pre-embedding unlabelled antibody PAP method. The protein is found in both fibrous and protoplasmic astrocytes and in the ependymal cells. The neurons, the oligodendrocytes as well as the microglial cells are lacking S-100. The labelled cells show a reaction product diffusely distributed in the cytoplasmic matrix and on specialized membranes, namely plasma membranes, outer mitochondrial membranes and membranes of the endoplasmic reticulum and Golgi apparatus. The astrocytic filaments and the axonemes of the ependymal cilia exhibit a strong immunoreactivity. The reaction product is also present in the nucleoplasm of the astrocytes and ependymal cells but it is absent from the nucleolus and nuclear envelope. This immunocytochemical data on tissue with satisfactory ultrastructural preservation, provides new information on the localization of the S-100 protein, and could contribute to the understanding of the biological role of the protein.     

Neurospecific protein S-100 in the synaptosomes of rat cerebral cortex]

Neurospecific S-100 protein was revealed by the methods of microelectrophoresis in the 15% polyacrylamide gel with a 0.1% sodium dodecylsulfate and by a highly purified S-100 protein "marker" in the composition of low molecular acidic proteins of the rat brain synaptosomes. The S-100 protein conten constitutes about 15-2o% of the low molecular acidic synaptosomal porteins in the rat brain.     

Immunochemical and immunohistochemical detection of S-100-like immunoreactivity in spinach tissues.

S-100 proteins represent a group of closely related acidic, calcium binding proteins originally isolated from the mammalian nervous system and later detected in non-neural cell types and in a wide variety of vertebrate and invertebrate species. The present study used immunochemical and immunohistochemical methods to extend the investigation of S-100 during phylogenesis to plant tissues. The presence of S-100-like immunoreactive material was detected in extracts of spinach (Spinacia oleracea L.) terminal buds and young leaves by the ELISA method and by Western blotting using different anti-S-100 rabbit antisera. Using the PAP method, serial sections of young spinach leaves treated with the same antisera exhibited an immunoreaction product that was confined to the cytoplasm and nucleus (but absent from the vacuoles) in meristematic, epidermal, and parenchymal cells. The present data enlarge the field of investigation of S-100 proteins in the search of the function(s) of S-100 in biological organisms.     

Spectroscopic studies on Tb3+ binding to S-100a protein.

Direct binding assay and fluorescence studies revealed that S-100a protein binds 2 mol of Tb3+/mol of protein at pH 6.6. The protein binds Tb3+ much more tightly than Ca2+, and the upper limit of the observed Kd value for Tb3+ is 3.5 x 10(-6) M. The Tb3+-binding site on the protein must be close to a tyrosine residue, as indicated by fluorescence excitation and emission spectra, where energy transfer from tyrosine is noted. Addition of Tb3+ resulted in a conformational change in the protein, as revealed by u.v.-difference spectroscopy and c.d. studies. Far-u.v. c.d. studies indicated the helical content to decrease from approx. 39% to 35% in the presence of Tb3+. From u.v.-difference-spectroscopy results the single tryptophan and the tyrosine chromophores in S-100a protein are blue-shifted (i.e. exposed to the solvent) in the presence of Tb3+ and the observed conformational changes are similar to those induced by Ca2+, suggesting that Tb3+ can be employed as a Ca2+ analogue in spectral studies with S-100a protein.     

Long-term adaptive response to dietary protein of hexose monophosphate shunt dehydrogenases in rat kidney tubules

We have studied the effects of several different macronutrients on the kinetic behaviour of rat renal glucose 6-phosphate dehydrogenase (G6PDH) and 6-phosphogluconate dehydrogenase (6PGDH). Rats were meal-fed with high-carbohydrate/low-protein, high-protein/low-carbohydrate and high-fat diets. High-protein increased renal G6PDH and 6PDGH activities by 66 per cent and 70 per cent respectively, without significantly changing the Km values of either and each Hexose monophosphate dehydrogenase activity increased steadily, reaching a significant difference on day 4. A rise in carbohydrate or fat in the diets, produced no significant change in either the activity or the kinetic parameters, Vmax and Km of the two dehydrogenases. In addition, the administration of a high-protein diet for 8 days significantly increased both the pentose phosphate pathway flux (92.6 per cent) and the kidney weigth (35 per cent), whereas no significant changes in these parameters were found when the animals were treated with the other diets. Our results suggest that an increase in the levels of dietary protein induces a rise in the intracellular levels of these enzymes. The possible role of this metabolic pathway in the kidneys under these nutritional conditions is also discussed.     

Predatory aggressiveness of the rat after intraventricular administration of individual brain-specific proteins of the S-100 group and their peptide fragments

Intraventricular injections to rats of the basic fraction of the brain specific protein S-100 in a concentration of 3 mg/ml, significantly facilitates the formation of their predatory aggression induced by the alimentary deprivation and social isolation, expressed in mice killing. This effect is not produced by fragments of S-100 molecules obtained as a result of treatment of the basic protein fraction by proteolytic enzymes. Administration of the minor S-100 fraction, albumin and rats summate brain proteins did not influence animals predatory aggression.