Multigenic factors associated with a hydrocephalus-like phenotype found in inter-subspecific consomic mouse strains

Hydrocephalus is a significant clinical condition in humans and is known to be a multifactorial neurologic disorder. It has been thought that genetic factors are closely involved in the etiology of congenital hydrocephalus, but further investigation is required to elucidate the genetic architecture of hydrocephalus. By analyzing breeding records of a panel of inter-subspecific consomic mouse strains, we found that consomic strains with MSM/Ms (MSM) chromosomes 4, 5, 7, 11, 15, and 17 showed a significantly higher incidence of hydrocephalus, whereas both parental strains, MSM and C57BL/6J (B6), rarely showed this abnormality. Further analysis of the consomic Chr 17 strain revealed that apparently normal individuals of this strain also exhibited increased brain ventricle size compared to B6 and had larger individual variation of ventricle size within the strain. Thus, we concluded that hydrocephalus is an extreme phenotype of individual ventricle size variation. We then established and analyzed several subconsomic strains of Chr 17 to identify genetic factors related to hydrocephalus-like phenotype and successfully mapped one genetic locus around the proximal region of Chr 17.     

Genetic heterogeneity in X-linked hydrocephalus: linkage to markers within Xq27.3.

X-linked hydrocephalus is a well-defined disorder which accounts for > or = 7% of hydrocephalus in males. Pathologically, the condition is characterized by stenosis or obliteration of the aqueduct of Sylvius. Previous genetic linkage studies have suggested the likelihood of genetic homogeneity for this condition, with close linkage to the DXS52 and F8C markers in Xq28. We have investigated a family with typical X-linked aqueductal stenosis, in which no linkage to these markers was present. In this family, close linkage was established to the DXS548 and FRAXA loci in Xq27.3. Our findings demonstrate that X-linked aqueductal stenosis may result from mutations at two different loci on the X chromosome. Caution is indicated in using linkage for the prenatal diagnosis of X-linked hydrocephalus.     

Impaired lymphatic cerebrospinal fluid absorption in a rat model of kaolin-induced communicating hydrocephalus

It has been assumed that the pathogenesis of hydrocephalus includes a cerebrospinal fluid (CSF) absorption deficit. Because a significant portion of CSF absorption occurs into extracranial lymphatics located in the olfactory turbinates, the purpose of this study was to determine whether CSF transport was compromised at this location in a kaolin-induced communicating (extraventricular) hydrocephalus model in rats. Under 1-3% halothane anesthesia, kaolin (n = 10) or saline (n = 9) was introduced into the basal cisterns of Sprague-Dawley rats, and the development of hydrocephalus was assessed 1 wk later using MRI. After injection of human serum albumin ((125)I-HSA) into a lateral ventricle, the tracer enrichment in the olfactory turbinates 30 min postinjection provided an estimate of CSF transport through the cribriform plate into nasal lymphatics. Lateral ventricular volumes in the kaolin group (0.073 +/- 0.014 ml) were significantly greater than those in the saline-injected animals (0.016 +/- 0.001 ml; P = 0.0014). The CSF tracer enrichment in the olfactory turbinates (expressed as percent injected/g tissue) in the kaolin rats averaged 0.99 +/- 0.39 and was significantly lower than that measured in the saline controls (5.86 +/- 0.32; P < 0.00001). The largest degree of ventriculomegaly was associated with the lowest levels of lymphatic CSF uptake with lateral ventricular expansion occurring only when almost all of the lymphatic CSF transport capacity had been compromised. We conclude that lymphatic CSF absorption is impaired in a kaolin-communicating hydrocephalus model and that the degree of this impediment may contribute to the severity of the induced disease.     

Protein-carbohydrate-lipid complex isolated from the cell envelopes of Chlamydia psittaci in alkaline buffer and ethylenediaminetetraacetate.

Exposure of isolated cell envelopes from purified infectious elementary (EB) of Chlamydia psittaci to sodium carbonate-bicarbonate buffer at pH 10 plus ethylenediaminetetraacetate (EDTA) results in partial solubilization of the total protein. The released materials represent 20% of the dry weight, 16% of the total protein, 40% of the total carbohydrate, and 9% of the total lipid of the cell envelopes. Sucrose density gradient centrifugation, and Sephadex G-200, Sepharose 4B, or diethylaminoethyl-cellulose column chromatography, reveal a protein-carbohydrate-lipid complex of several hundred thousand molecular weight that contains 50% protein. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis of the isolated EB cell envelopes reveals two major protein bands, A and B, with estimated molecular masses of approximately 85,000 and 53,000, respectively, both of which also stain for the presence of carbohydrate and lipid. Gel electrophoresis of the protein-carbohydrate-lipid complex reveals two protein bands, C and D, with estimated molecular weights of approximately 17,000 and 13,000, respectively, which contain lipid and a small amount of carbohydrate; bands A and B are not present in the complex. Gel electrophoresis of the cell envelope residues after extraction of the complex with alkali and EDTA shows a single main band, corresponding to the position of band B, which contains protein, carbohydrate, and lipid; band A is completely missing. B and A is believed to be a component of the complex, which is split into two subunits on alkali solubilization.     

Variations of prolactin content in human cerebrospinal fluid after metoclopramide and morphine

Serum and cerebrospinal fluid (CSF) prolactin (PRL) concentrations were determined in fourteen patients of both sexes suffering from hydrocephalus, in basal conditions and after i.m. administration of 10 mg metoclopramide or 10 mg morphine. A significant increase in both serum and CSF hormone values was found after administration of both drugs. Serum and CSF PRL values after metoclopramide administration increased earlier and to a greater extent than after morphine. Furthermore, the metoclopramide induced CSF PRL increase immediately followed the serum peak, whereas after morphine administration an evident delay in the CSF hormone peak with respect to the serum increase was found. These data suggest that PRL entry in the CSF compartment is subject to a controlling mechanism which acts at the blood/brain barrier.     

Reactive microglia and mitochondrial unfolded protein response following ventriculomegaly and behavior defects in kaolin-induced hydrocephalus

Ventriculomegaly induced by the abnormal accumulation of cerebrospinal fluid (CSF) leads to hydrocephalus, which is accompanied by neuroinflammation and mitochondrial oxidative stress. The mitochondrial stress activates mitochondrial unfolded protein response (UPRmt), which is essential for mitochondrial protein homeostasis. However, the association of inflammatory response and UPRmt in the pathogenesis of hydrocephalus is still unclear. To assess their relevance in the pathogenesis of hydrocephalus, we established a kaolin-induced hydrocephalus model in 8-week-old male C57BL/6J mice and evaluated it over time. We found that kaolin-injected mice showed prominent ventricular dilation, motor behavior defects at the 3-day, followed by the activation of microglia and UPRmt in the motor cortex at the 5-day. In addition, PARP-1/NF-κB signaling and apoptotic cell death appeared at the 5-day. Taken together, our findings demonstrate that activation of microglia and UPRmt occurs after hydrocephalic ventricular expansion and behavioral abnormal-ities which could be lead to apoptotic neuronal cell death, providing a new perspective on the pathogenic mechanism of hydrocephalus.     

Strain difference of the mouse in manifestation of hydrocephalus following prenatal methylmercury exposure

Genetic background may influence susceptibility to hydrocephalus. In the present experiment we compared the manifestation of hydrocephalus following prenatal methylmercury exposure among strains of mice which sporadically develop or never develop spontaneous hydrocephalus. Pregnant mice of the B10.D2 congenic strain were given a single oral dose of 10 mg/kg methylmercuric chloride on one of days 14 through 17 of pregnancy and allowed to give birth and rear their litters. The incidence of grossly apparent hydrocephalus in the offspring at 30 days of age following treatment on day 14, 15, 16, or 17 of pregnancy was 67, 88, 75, and 48%, respectively; that of sham-treated and untreated offspring was 5 and 4%, respectively. In addition, there were some brains showing slight dilatation of the lateral ventricles. Pregnant females of C57BL/10 (B10) or DBA/2 (D2) strain were also treated with 10 mg/kg methylmercury on day 15 of pregnancy. The incidence of hydrocephalus at 30 days of age in untreated and dosed B10 mice was 0.8 and 54%, respectively. Hydrocephalus failed to develop in D2 mice. The hydrocephalus is a communicating type. Occlusion of the cerebral aqueduct with glial reaction and caudal displacement of the cerebellum are considered to be secondary changes. The results indicate that the susceptibility to methylmercury-induced hydrocephalus is under genetic control in mice.     

Report of a collaborative study to assess the suitability of a reference reagent for antibodies to hepatitis E virus.

Several commercial and "in-house" assays have been developed for the detection of antibodies to hepatitis E virus, a major causative agent of enterically transmitted non-A non-B hepatitis. As these kits contain a variety of synthetic peptides or recombinant proteins, greater standardisation is required. A collaborative study was therefore carried out to assess the suitability of a freeze dried preparation designated 95/584 to serve as a reference reagent for hepatitis E virus serum IgG. Preparation 95/584, which is a serum from a previously infected individual, was assayed along with four coded samples, one of which D, was a coded duplicate of 95/584, and three individual sera, coded A, B and C. These preparations were sent to seven laboratories in five countries who tested them in eight different enzyme immunoassays. In most laboratories the coded duplicate gave a mean potency of within 20% of the candidate reference reagent despite the wide range of assays used. However, the potencies of the coded samples which were from different individuals gave somewhat variable potencies relative to the candidate reference reagent. This is not surprising as each sample will have varying proportions of antibodies against individual viral proteins and result in the variation in results observed. Nevertheless, this material will be of use in the standardisation of diagnostic tests for use in sero-prevalence studies and for assessing immunity. Preparation 95/584 was found to be suitable to serve as a reference reagent for hepatitis E serum IgG and has been established as an interim Reference Reagent for Human anti-hepatitis E serum. Each ampoule contains 50 Units per ampoule.     

Normal-pressure hydrocephalus due to tentorial meningioma.

Normal-pressure hydrocephalus and the associated triad of dementia, apraxic gait and urinary incontinence may be casued by various, sometimes unsuspected, lesions, usually those that block the flow of cerebrospinal fluid (CSF) around the tentorium. A 58-year-old woman with insidious onset of behaviour and gait problems had occult, normal-pressure hydrocephalus and a tentorial meningioma, resection of which produced complete recovery. This case demonstrates that a distinction must be made between the syndrome of normal-pressure hydrocephalus resulting from disturbance in CSF dynamics and the pathophysiologic features of the underlying lesion.     

Purification and characterization of three forms of glutathione S-transferase A. A comparative study of the major YaYa-, YbYb- and YcYc-containing glutathione S-transferases.

Rat liver glutathione S-transferases have previously been defined by their elution behaviour from DEAE-cellulose and CM-cellulose as M, E, D, C, B, A and AA. These enzymes are dimeric proteins which comprise subunits of mol.wt. 22 000 (Ya), 23 500 (Yb) or 25 000 (Yc). Evidence is presented that YaYa protein, one of two previously described lithocholate-binding proteins which exhibit transferase activity, is an additional enzyme which is not included in the M, E, D, C, B, A and AA nomenclature. We therefore propose that this enzyme is designated transferase YaYa. Transferases YaYa, C, A and AA have molecular weights of 44 000, 47 000, 47 000 and 50 000 respectively and each comprises two subunits of identical size. These enzymes were purified to allow a study of their structural and functional relationships. In addition, transferase A was further resolved into three forms (A1, A2 and A3) which possess identical activities and structures and appear to be the product of a single gene. Transferases YaYa, C, A and AA each had distinct enzymic properties and were inhibited by cholate. The recently proposed proteolytic model, which attributes the presence of multiple forms of glutathione S-transferase activity to partial proteolysis of transferase AA, was tested and shown to be highly improbable. Peptide maps showed significant differences between transferases YaYa, C, A and AA. Immunotitration studies demonstrated that antisera raised against transferases YaYa and C did not precipitate transferase AA.     

Presence of a novel nuclear lamina protein in Raji cells.

In mammalian tissues, the nuclear lamina is composed of the major lamins A, B, and C, and minor lamins D/E. Although lamin B is present in all cell types, lamins A and C are absent from embryonic cells and most undifferentiated cells from hematopoietic lineage. We have investigated the nuclear lamina protein composition of the Raji cell line, lymphoblast-like cells established from a Burkitt lymphoma patient. Lamins A and C were confirmed absent by immunodetection and Northern blot analysis. Besides lamins B and D/E, a protein migrating around 71 kilodaltons was recognized by a serum directed against the nuclear lamina of BHK-21 fibroblasts. Cellular localization by sequential extraction established this 71-kilodalton protein as an exclusive component of the nuclear lamina fraction. These results indicate that the nuclear lamina has a more complex composition than previously thought to be the case for cells devoid of lamins A and C.     

Steroid-independent male sexual behavior in B6D2F2 male mice

It is well established that male sexual behavior (MSB) is regulated by gonadal steroids; however, individual differences in MSB, independent of gonadal steroids, are prevalent across a wide range of species, and further investigation is necessary to advance our understanding of steroid-independent MSB. Studies utilizing B6D2F1 hybrid male mice in which a significant proportion retain MSB after long-term orchidectomy, identified as steroid-independent-maters (SI-maters), have begun to unravel the genetic underpinnings of steroid-independent MSB. A recent study demonstrated that steroid-independent MSB is a heritable behavioral phenotype that is mainly passed down from B6D2F1 hybrid SI-maters when crossed with C57BL6J female mice. To begin to uncover whether the strain of the dam plays a role in the inheritance of steroid-independent MSB, B6D2F1 hybrid females were crossed with B6D2F1 hybrid males. While the present study confirms the finding that steroid-independent MSB is a heritable behavioral phenotype and that SI-mater sires are more likely to pass down some components of MSB than SI-non-maters to their offspring, it also reveals that the B6D2F2 male offspring that were identified as SI-maters that displayed the full repertoire of steroid-independent MSB had the same probability of being sired from either a B6D2F1 SI-mater or SI-non-mater. These results, in conjunction with previous findings, indicate that the specific chromosomal loci pattern that codes for steroid-independent MSB in the B6D2F2 male offspring may result regardless of whether the father was a SI-mater or SI-non-mater, and that the maternal strain may be an important factor in the inheritance of steroid-independent MSB.     

Cloning of the rfb gene cluster of a group C2 Salmonella strain: comparison with the rfb regions of groups B and D

We report the cloning and mapping of the entire rfb gene cluster of a group C2 Salmonella strain. Comparison with the rfb region of group B strain LT2 and group D strain Ty2 reveals an 11.8 kb central region of limited similarity flanked by regions of high similarity. The genes from the central region confer a group C2 O-antigen structure on a Salmonella LT2 partial delete strain. The significance of this region in relation to function and evolutionary origin is discussed. We also report evidence for the existence of an O-antigen chain-length determinant in Escherichia coli K12 and propose a model for a possible mechanism by which a preferred chain length is determined.     

High-pressure hydrocephalus: a novel analytical modeling approach

Hydrocephalus is an abnormal accumulation of cerebrospinal fluid (CSF) within ventricles and subarachnoid space (SAS) as a result of disturbances in secretion or absorption procedures. It is believed that arachnoid villi cells, which are microscopic projections of pia-arachnoid mater that extend into venous channels in sagittal sinus, are the main sites for CSF absorption, but it is tempting to speculate that a significant portion of CSF is removed from the SAS by nasal lymphatic vessels around olfactory nerve. Thus, in this paper, we propose an analytical model of CSF-lymphatic-blood circulation, in which these two output pathways for CSF absorption have been considered. Mathematical relations governing the pressures in different interacting compartments of the brain are considered. In addition, for increasing the similarity of our model to the physiological conditions, the bulk flow mechanism, which is supposed to occur during CSF absorption, has been considered in our model. We used our model to simulate hydrocephalus. The results indicate that the lymphatic disorders have more considerable effect in decreasing CSF absorption, compared to the disturbances in arachnoid villi cells. Based on our modeling, we believe that disorders in lymphatic pathway may be a cause of high-pressure hydrocephalus. Surely experimental studies are required to validate our hypothesis.     

Junctional Adhesion Molecule (JAM)-C Deficient C57BL/6 Mice Develop a Severe Hydrocephalus

The junctional adhesion molecule (JAM)-C is a widely expressed adhesion molecule regulating cell adhesion, cell polarity and inflammation. JAM-C expression and function in the central nervous system (CNS) has been poorly characterized to date. Here we show that JAM-C^−/− mice backcrossed onto the C57BL/6 genetic background developed a severe hydrocephalus. An in depth immunohistochemical study revealed specific immunostaining for JAM-C in vascular endothelial cells in the CNS parenchyma, the meninges and in the choroid plexus of healthy C57BL/6 mice. Additional JAM-C immunostaining was detected on ependymal cells lining the ventricles and on choroid plexus epithelial cells. Despite the presence of hemorrhages in the brains of JAM-C^−/− mice, our study demonstrates that development of the hydrocephalus was not due to a vascular function of JAM-C as endothelial re-expression of JAM-C failed to rescue the hydrocephalus phenotype of JAM-C^−/− C57BL/6 mice. Evaluation of cerebrospinal fluid (CSF) circulation within the ventricular system of JAM-C^−/− mice excluded occlusion of the cerebral aqueduct as the cause of hydrocephalus development but showed the acquisition of a block or reduction of CSF drainage from the lateral to the 3^rd ventricle in JAM-C^−/− C57BL/6 mice. Taken together, our study suggests that JAM-C^−/− C57BL/6 mice model the important role for JAM-C in brain development and CSF homeostasis as recently observed in humans with a loss-of-function mutation in JAM-C.     

Dopamine D1B Receptor Chimeras Reveal Modulation of Partial Agonist Activity by Carboxyl-Terminal Tail Sequences

Abstract: NNC 01-0012, a second-generation benzazepine compound, pharmacologically differentiates multiple vertebrate D1 receptor subtypes (D1A, D1B, D1C, and D1D) and displays high selectivity and affinity for dopamine D1C receptors. Functionally, whereas NNC 01-0012 acts as a full or poor antagonist at D1C and D1A receptor-mediated cyclic AMP production, respectively, it exhibits partial agonist activity at the D1B receptor. To define some of the structural motifs that regulate the pharmacological and functional differentiation of vertebrate dopamine D1 receptors by NNC 01-0012, a series of receptor chimeras were constructed in which the divergent carboxyl-terminal (CT) receptor tails were replaced with the corresponding sequences of D1A, D1B, or D1C receptors. Substitution of the vertebrate D1B carboxyl-terminal-tail at position Tyr³⁴⁵ with carboxyl-terminal-tail sequences of the D1A receptor abolished the partial agonist activity of NNC 01-0012 without affecting dopamine-stimulated cyclic AMP accumulation. At vertebrate D1B/D1Cct -tail receptor mutants, however, the intrinsic activity of the partial agonist NNC 01-0012 (10 µM) was markedly enhanced (～60% relative to 10 µM dopamine) with no concomitant alteration in the molecule's ligand binding affinity or constitutive activity of the chimeric receptor. Similar results were obtained with other benzazepines such as SKF-38393 and SCH-23390, which act as partial agonists at vertebrate D1B receptors. Substitution of D1A and D1C receptor carboxyl-terminal tails with sequences encoded by the D1B receptor carboxyl-terminal tail did not, however, produce receptors with functional characteristics significantly different from wild type. Taken together, these data clearly suggest that in addition to well-characterized domains and amino acid residues in the third cytoplasmic loop, partial agonist activity at the D1B receptor is modulated by sequence-specific motifs within the carboxyl-terminal tail, a region that may underlie the possible structural basis for functionally divergent roles of multiple dopamine D1-like receptors.     

Congenital hydrocephalus caused by exposure to low level X-radiation at early gestational stage in mice.

Hydrocephalus is a severe and often lethal birth defect in humans resulting from excess accumulation off cerebrospinal fluid (CSF) in the cranial vault, accompanied by enlargement of the head, prominence of the forehead and atrophy of the brain. A wide variety of teratogenic procedures have been used to obtain congenital hydrocephalus in laboratory animals. Radiation, infections, trypan blue, hypervitaminosis A, salisylates and nutritional deficiencies were considered as the teratogens. Several investigators induced congenital hydrocephalus following irradiation of pregnant animals. The purpose off this study is to elucidate the pathogenesis off congenital hydrocephalus induced by X-irradiation in mice. Since the sensitive period for malformations produced by X-radiation in mice ranges from gestational day 7 (G7) to Gl3.     

Alternative substrate and inhibition kinetics of aminoglycoside nucleotidyltransferase 2'-I in support of a Theorell-Chance kinetic mechanism

Aminoglycoside nucleotidyltransferase 2'-I conveys multiple antibiotic resistance to Gram-negative bacteria because the enzyme adenylylates a broad range of aminoglycoside antibiotics as substrates [Gates, C. A., & Northrop, D. B. (1988) Biochemistry (preceding paper in this issue)]. The enzyme also catalyzes the transfer of a variety of nucleotides [Van Pelt, J. E., & Northrop, D. B. (1984) Arch. Biochem. Biophys. 230, 250-263]. This doubly broad substrate specificity makes it an excellent candidate for application of the alternative substrate diagnostic [Radika, K., & Northrop, D. B. (1984) Anal. Biochem. 141, 413-417] as a means to determine its kinetic mechanism. The kinetic patterns presented here are composed of one set of intersecting lines and one coincident line and are consistent with a Theorell-Chance kinetic mechanism in which nucleotide binding precedes aminoglycosides, pyrophosphate is released prior to the nucleotidylated aminoglycoside (Q), and turnover is controlled by the rate-limiting release of the final product. Substrate inhibition by tobramycin (B) is partial and uncompetitive versus Mg-ATP, indicating that B binds to the EQ complex, but not in the usual dead-end fashion common to an ordered sequential release of products; instead, Q may escape from the abortive EQB complex at a finite rate. Dead-end inhibition by neomycin C (I) is also partial and uncompetitive versus Mg-ATP but is slope-linear, intercept-hyperbolic, partial noncompetitive versus gentamicin A; both kinetic patterns signify the formation of a partial abortive EQI complex.(ABSTRACT TRUNCATED AT 250 WORDS)