Regulation by commensal bacteria of neurogenesis in the subventricular zone of adult mouse brain.

In the mouse olfactory bulb (OB), interneurons such as granule cells and periglomerular cells are continuously replaced by adult-born neurons, which are generated in the subventricular zone (SVZ) of the brain. We have now investigated the role of commensal bacteria in regulation of such neuronal cell turnover in the adult mouse brain. Administration of mixture of antibiotics to specific pathogen-free (SPF) mice markedly attenuated the incorporation of bromodeoxyuridine (BrdU) into the SVZ cells. The treatment with antibiotics also reduced newly generated BrdU-positive neurons in the mouse OB. In addition, the incorporation of BrdU into the SVZ cells of germ-free (GF) mice was markedly reduced compared to that apparent for SPF mice. In contrast, the reduced incorporation of BrdU into the SVZ cells of GF mice was recovered by their co-housing with SPF mice, suggesting that commensal bacteria promote the incorporation of BrdU into the SVZ cells. Finally, we found that administration of ampicillin markedly attenuated the incorporation of BrdU into the SVZ cells of SPF mice. Our results thus suggest that ampicillin-sensitive commensal bacteria regulate the neurogenesis in the SVZ of adult mouse brain.

Central nervous system innervation of the penis as revealed by the transneuronal transport of pseudorabies virus.

Transneuronal tracing techniques were used in order to identify putative spinal interneurons and brainstem sites involved in the control of penile function. Pseudorabies virus was injected into the corpus cavernosus tissue of the penis in rats. After a four day survival period, rats were perfused with fixative and virus-labelled neurons were identified by immunohistochemistry. Postganglionic neurons were retrogradely labelled in the major pelvic ganglia. In the spinal cord, sympathetic and parasympathetic preganglionic neurons were labelled transneuronally. Presumptive interneurons were also labelled in the lower thoracic and lumbosacral spinal cord in locations consistent with what is currently known about such interneurons. In the brainstem, transneuronally labelled neurons were found in the medulla, pons and hypothalamus. Regions consistently labelled included the nucleus paragigantocellularis, parapyramidal reticular formation of the medulla, raphe pallidus, raphe magnus, A5 noradrenergic cell group, Barrington's nucleus and the paraventricular nucleus of the hypothalamus. This study confirmed previous studies from our lab and others concerning the preganglionic and postganglionic neurons innervating the penis. The number, morphology and location of these neurons were consistent with labelling seen following injection of conventional tracers into the penis. The brainstem nuclei labelled in this study were also consistent with what is currently known about the brainstem control of penile function. The labelling appeared to be highly specific, in that descending systems involved in other functions were not labelled. These results provide further evidence that the pseudorabies virus transneuronal tracing technique is a valuable method for identifying neural circuits mediating specific functions.

Transneuronal labeling of spinal interneurons and sympathetic preganglionic neurons after pseudorabies virus injections in the rat medial gastrocnemius muscle.

The distribution of retrogradely and transneuronally labeled neurons was studied in CNS of rats 4 days after injections of the Bartha strain of pseudorabies virus (PRV) into the medial gastrocnemius (MG) muscle. Tissue sections were processed for immunohistochemical detection of PRV. Retrogradely labeled cells were identified in the ipsilateral MG motor column in the caudal L4 and the L5 spinal segments. In order to evaluate the efficacy of PRV retrograde cell body labeling, the number of PRV retrogradely labeled neurons in the MG motor column was compared to the number labeled with two conventional retrograde cell body markers--Fluoro-Gold and cholera toxin-HRP. A ratio of 1:3 representing medium-sized (less than 30 microns) versus large neurons (greater than 30 microns) was found in the Fluoro-Gold dye experiments; a 1:2 ratio was seen in the PRV experiments. In contrast, when cholera toxin-HRP was used as a retrograde marker, mainly large neurons were labeled; the medium-to-large cell body ratio was 1:10 suggesting cholera toxin-HRP may have a greater affinity for the terminals of alpha-motoneurons as opposed to gamma-motoneurons. Transneuronally labeled cells were identified in the L1-L6 spinal gray matter, intermediolateral cell column (T11-L2), lateral spinal nucleus and medial part of lamina VII in C4 and C5 spinal segments, brainstem (caudal raphe nuclei, rostral ventrolateral medulla, A5 cell group, paralemniscal nucleus, locus coeruleus, subcoeruleus nucleus, red nucleus) and paraventricular hypothalamic nucleus. In the L5 spinal cord, transneuronally labeled neurons were seen in the ipsilateral spinal laminae I and II and bilaterally in spinal laminae IV-VIII, and X. Similar results were obtained in rats that had chronic unilateral L3-L6 dorsal rhizotomies indicating most of the labeling was due to retrograde transneuronal cell body labeling. In order to determine whether PRV was transported into the spinal cord by the dorsal root axons, the ipsilateral dorsal root ganglia (DRGs) were examined for PRV immunoreactivity; none was found. However, using the polymerase chain reaction, viral DNA was shown to be present in the ipsilateral DRGs indicating that some of spinal cord cell body labeling may have resulted from anterograde transneuronal labeling, as well.