Catecholaminergic neurons are affected in several neurological and psychiatric diseases. Tyrosine hydroxylase (TH) is the first, rate-limiting enzyme in catecholamine synthesis. We report a knockin mouse expressing Cre-recombinase from the 3'-untranslated region of the endogenous Th gene by means of an internal ribosomal entry sequence (IRES). The resulting Cre expression matches the normal pattern of TH expression, while the pattern and level of TH are not altered in the knockin mouse. Crossings with two different LacZ reporter mice demonstrated Cre-mediated genomic recombination in TH expressing tissues. In addition, LacZ was found in some unexpected cell populations (including oocytes), indicating recombination due to transient developmental TH expression. Our novel knockin mouse can be used for generation of tissue-specific or general knockouts (depending on scheme of crossing) in mice carrying genes flanked by loxP sites. This knockin mouse can also be used for tracing cell lineages expressing TH during development.

Cerebral gene expressions change in response to traumatic brain injury (TBI), and future trauma treatment may improve with increased knowledge about these regulations. We subjected C57BL/6J mice to injury by controlled cortical impact (CCI). At various time points post-injury, mRNA from neocortex and hippocampus was isolated, and transcriptional alterations studied using quantitative real-time polymerase chain reaction (PCR) and gene array analysis. Spatial distribution of enhanced expression was characterized by in situ hybridization. Products of the upregulated transcripts serve functions in a range of cellular mechanisms, including stress, inflammation and immune responses, and tissue remodeling. We also identified increased transcript levels characterizing reactive astrocytes, oligodendrocytes, and microglia, and furthermore, we demonstrated a novel pattern of scattered cell clusters expressing the chemokine Cxcl10. Notably, a sustained increase in integrin alpha X (Itgax), characterizing antigen-presenting dendritic cells, was found with the transcript located to similar cell clusters. In contrast, T-cell receptor alpha transcript showed only a modest increase. The induced P-selectin (Selp) expression level in endothelial cells, and chemokines from microglia, may guide perivascular accumulation of extravasating inflammatory monocytes differentiating into dendritic cells. In conclusion, our study shows that following TBI, secondary injury chiefly involves inflammatory processes and chemokine signaling, which comprise putative targets for pharmaceutical neuroprotection.

Brain trauma is known to activate inflammatory cells via various chemokine signals although their interactions remain to be characterized. Mice deficient in Ccl3, Ccr2 or Cxcl10 were compared with wildtype mice after controlled cortical impact injury. Expression of Ccl3 in wildtypes was rapidly upregulated in resident, regularly spaced reactive microglia. Ccl3-deficiency enhanced endothelial expression of platelet selectin and invasion of peripheral inflammatory cells. Appearance of Ccr2 transcripts, encoding the Ccl2 receptor, reflected invasion of lysozyme 2-expressing phagocytes and classical antigen-presenting dendritic cells expressing major histocompatibility complex class II. Ccr2 also directed clustered plasmacytoid dendritic cells positive for the T-cell attracting chemokine Cxcl10. A reduction in Ccr2 and dendritic cells was found in injured wildtype cortex after cyclophosphamide treatment resembling effects of Ccr2-deficiency. The findings demonstrate the feasibility to control inflammation in the injured brain by regulating chemokine-dependent pathways.

Cerebral gene expression changes in response to traumatic brain injury will provide useful information in the search for future trauma treatment. In order to characterize the outcome of mild brain injury, we studied C57BL/6J mice in a weight-drop, closed head injury model. At various times post-injury, mRNA was isolated from neocortex and hippocampus and transcriptional alterations were studied using quantitative reverse transcriptase PCR and gene array analysis. At three days post-injury, the results showed unilateral injury responses, both in neocortex and hippocampus, with the main effect seen on the side of the skull hit by the dropping weight. Upregulated transcripts encoded products characterizing reactive astrocytes, phagocytes, microglia, and immune-reactive cells. Markers for oligodendrocytes and T-cells were not altered. Notably, strong differences in the responses among individual mice were seen (e.g., for the Gfap transcript expressed by reactive astrocytes and the chemokine Ccl3 transcript expressed by activated microglial cells). In conclusion, mild TBI chiefly activates transcripts leading to tissue signaling, inflammatory processes, and chemokine signaling, as in focal brain injury, suggesting putative targets for drug development.

Traumatic brain injury (TBI) in the mouse results in the rapid appearance of scattered clusters of cells expressing the chemokine Cxcl10 in cortical and subcortical areas. To extend the observation of this unique pattern, we used neuropathological mouse models using quantitative reverse transcriptase-polymerase chain reaction, gene array analysis, in-situ hybridization and flow cytometry. As for TBI, cell clusters of 150-200 mum expressing Cxcl10 characterize the cerebral cortex of mice carrying a transgene encoding the Swedish mutation of amyloid precursor protein, a model of amyloid Alzheimer pathology. The same pattern was found in experimental autoimmune encephalomyelitis in mice modelling multiple sclerosis. In contrast, mice carrying a SOD1(G93A) mutant mimicking amyotrophic lateral sclerosis pathology lacked such cell clusters in the cerebral cortex, whereas clusters appeared in the brainstem and spinal cord. Mice homozygous for a null mutation of the Cxcl10 gene did not show detectable levels of Cxcl10 transcript after TBI, confirming the quantitative reverse transcriptase-polymerase chain reaction and in-situ hybridization signals. Moreover, unbiased microarray expression analysis showed that Cxcl10 was among 112 transcripts in the neocortex upregulated at least threefold in both TBI and ageing TgSwe mice, many of them involved in inflammation. The identity of the Cxcl10(+) cells remains unclear but flow cytometry showed increased numbers of activated microglia/macrophages as well as myeloid dendritic cells in the TBI and experimental autoimmune encephalomyelitis models. It is concluded that the Cxcl10(+) cells appear in the inflamed central nervous system and may represent a novel population of cells that it may be possible to target pharmacologically in a broad range of neurodegenerative conditions.

mRNA for bone morphogenetic protein receptor type II (BMPR-II) was mapped to different neurons in peripheral ganglia and spinal cord of the chicken embryo. The expression of this serine/threonine kinase receptor partially overlaps with that of tyrosine kinase receptors Trk and Ret. Biological activities of osteogenic protein-1 (OP-1), a documented ligand for BMPR-II, were tested in explanted embryonic chicken ganglia and dissociated ganglionic neurons. OP-1 had only a limited stimulatory effect on neuronal survival. However, OP-1 combined with either neurotrophin-3 (NT-3, a relative of nerve growth factor) or glial cell line-derived neurotrophic factor (GDNF) potentiated neuronal survival three- to fourfold. We also show that OP-1 strongly potentiates nerve fiber outgrowth from ganglia stimulated with NT-3 or GDNF. Signaling by BMPR-II in neurons may potentiate the tyrosine kinase pathway activated by NT-3 and GDNF. The data suggest that morphogenetic proteins may modulate neurotrophic activities during neuronal development and plasticity.

Cerebral gene expression changes in response to traumatic brain injury will provide useful information in the search for future trauma treatment. In order to characterize the outcome of mild brain injury, we studied C57BL/6J mice in a weight-drop, closed head injury model. At various times post-injury, mRNA was isolated from neocortex and hippocampus and transcriptional alterations were studied using quantitative reverse transcriptase PCR and gene array analysis. At three days post-injury, the results showed unilateral injury responses, both in neocortex and hippocampus, with the main effect seen on the side of the skull hit by the dropping weight. Upregulated transcripts encoded products characterizing reactive astrocytes, phagocytes, microglia, and immune-reactive cells. Markers for oligodendrocytes and T-cells were not altered. Notably, strong differences in the responses among individual mice were seen (e.g., for the Gfap transcript expressed by reactive astrocytes and the chemokine Ccl3 transcript expressed by activated microglial cells). In conclusion, mild TBI chiefly activates transcripts leading to tissue signaling, inflammatory processes, and chemokine signaling, as in focal brain injury, suggesting putative targets for drug development.

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