Complement system activation contributes to the ependymal damage induced by microbial neuraminidase

Background In the rat brain, a single intracerebroventricular injection of neuraminidase from Clostridium perfringens induces ependymal detachment and death. This injury occurs before the infiltration of inflammatory blood cells; some reports implicate the complement system as a cause of these injuries. Here, we set out to test the role of complement. Methods The assembly of the complement membrane attack complex on the ependymal epithelium of rats injected with neuraminidase was analyzed by immunohistochemistry. Complement activation, triggered by neuraminidase, and the participation of different activation pathways were analyzed by Western blot. In vitro studies used primary cultures of ependymal cells and explants of the septal ventricular wall. In these models, ependymal cells were exposed to neuraminidase in the presence or absence of complement, and their viability was assessed by observing beating of cilia or by trypan blue staining. The role of complement in ependymal damage induced by neuraminidase was analyzed in vivo in two rat models of complement blockade: systemic inhibition of C5 by using a function blocking antibody and testing in C6-deficient rats. Results The complement membrane attack complex immunolocalized on the ependymal surface in rats injected intracerebroventricularly with neuraminidase. C3 activation fragments were found in serum and cerebrospinal fluid of rats treated with neuraminidase, suggesting that neuraminidase itself activates complement. In ventricular wall explants and isolated ependymal cells, treatment with neuraminidase alone induced ependymal cell death; however, the addition of complement caused increased cell death and disorganization of the ependymal epithelium. In rats treated with anti-C5 and in C6-deficient rats, intracerebroventricular injection of neuraminidase provoked reduced ependymal alterations compared to non-treated or control rats. Immunohistochemistry confirmed the absence of membrane attack complex on the ependymal surfaces of neuraminidase-exposed rats treated with anti-C5 or deficient in C6. Conclusions These results demonstrate that the complement system contributes to ependymal damage and death caused by neuraminidase. However, neuraminidase alone can induce moderate ependymal damage without the aid of complement

Distribution of diacylglycerol lipase alpha, an endocannabinoid synthesizing enzyme, in the rat forebrain

1,2-diacylglycerol lipase alpha (DAGLα) is responsible for the biosynthesis and release of 2-arachidonoyl-glycerol (2-AG), the most abundant endocannabinoid in the brain. Although its expression has been detected in discrete regions, we showed here an integrated description of the distribution of DAGLα mRNA and protein in the rat forebrain using in situ hybridization histochemistry and immunohistochemistry. As novelty, we described the distribution of DAGLα protein expression in the olfactory system, the rostral migratory stream, neocortex, septum, thalamus, and hypothalamus. Similar DAGLα immunostaining pattern was also found in the brain of wild-type, but not of DAGLα knockout mice. Immunohistochemical data were correlated by the identification of DAGLα mRNA expression, for instance, in the somata of specific cells in olfactory structures, rostral migratory stream and neocortex, cells in some septal-basal-amygdaloid areas and the medial habenula, and magnocellular cells of the paraventricular hypothalamic nucleus. This widespread neuronal distribution of DAGLα is consistent with multiple roles for endocannabinoids in synaptic plasticity, including presynaptic inhibition of neurotransmitter release. We discuss our comparative analysis of the forebrain expression patterns of DAGLα and other components of the endocannabinoid signaling system, including the CB1 receptor, monoacylglyceride lipase (MAGL), and fatty acid amide hydrolase (FAAH), providing some insight into the potential physiological and behavioral roles of this system. © 2011 IBRO.Peer Reviewe

Neurogenesis in explants from the walls of the lateral ventricle of adult bovine brain: Role of endogenous IGF-1 as a survival factor

Previous studies have shown the existence of proliferating cells in explants from bovine (Bos Taurus) lateral ventricle walls that were maintained for several days in vitro in the absence of serum and growth factors. In this study we have characterized the nature of new cells and have assessed whether the insulin-like growth factor-1 (IGF-1) receptor regulates their survival and/or proliferation. The explants were composed of the ependymal layer and attached subependymal cells. Ependymal cells in culture were labelled with glial markers (S-100, vimentin, GFAP, BLBP, 3A7 and 3CB2) and did not incorporate bromodeoxiuridine when this molecule was added to the culture media. Most subependymal cells were immunoreactive for βIII-tubulin, a neuronal marker, and did incorporate bromodeoxiuridine. Subependymal neurons displayed immunoreactivity for IGF-1 and its receptor and expressed IGF-1 mRNA, indicating that IGF-1 is produced in the explants and may act on new neurons. Addition to the culture media of an IGF-1 receptor antagonist, the peptide JB1, did not affect the incorporation of bromodeoxiuridine to proliferating subependymal cells. However, JB1 significantly increased the number of TUNEL positive cells in the subependymal zone, suggesting that IGF-1 receptor is involved in the survival of subependymal neurons. In conclusion, these findings indicate that neurogenesis is maintained in explants from the lateral cerebral ventricle of adult bovine brains and that IGF-1 is locally produced in the explants and may regulate the survival of the proliferating neurons.Peer Reviewe