Intermittent hypoxia causes a form of serotonin-dependent synaptic plasticity in the spinal cord known as phrenic long-term facilitation (pLTF). Here we show that increased synthesis of brain-derived neurotrophic factor (BDNF) in the spinal cord is necessary and sufficient for pLTF in adult rats. We found that intermittent hypoxia elicited serotonin-dependent increases in BDNF synthesis in ventral spinal segments containing the phrenic nucleus, and the magnitude of these BDNF increases correlated with pLTF magnitude. We used RNA interference (RNAi) to interfere with BDNF expression, and tyrosine kinase receptor inhibition to block BDNF signaling. These disruptions blocked pLTF, whereas intrathecal injection of BDNF elicited an effect similar to pLTF. Our findings demonstrate new roles and regulatory mechanisms for BDNF in the spinal cord and suggest new therapeutic strategies for treating breathing disorders such as respiratory insufficiency after spinal injury. These experiments also illustrate the potential use of RNAi to investigate functional consequences of gene expression in the mammalian nervous system in vivo.

Although microglial activation is associated with all CNS disorders, many of which are sexually dimorphic or age-dependent, little is known about whether microglial basal gene expression is altered with age in the healthy CNS or if it is sex-dependent. Analysis of microglia from the brains of 3 day (P3) - to 12 month-old male and female C57Bl/6 mice revealed distinct gene expression profiles during postnatal development that differ significantly from adulthood. Microglia at P3 are characterized by relatively high iNOS, TNFα and arginase-1 mRNA levels, whereas P21 microglia have increased expression of CD11b, TLR4 and FcRγI. Adult microglia (2-4 months) are characterized by low pro-inflammatory cytokine expression that increases by 12 months of age. Age-dependent differences in microglial gene expression suggest that microglia likely undergo phenotypic changes during ontogenesis, although in the healthy brain they did not exclusively express either M1 or M2 phenotypic markers at any time. Interestingly, microglia were sexually dimorphic only at P3 when females had higher expression of inflammatory cytokines than males, although there were no sex differences in estrogen receptor expression at this or any other time evaluated here. Compared to microglia in vivo, primary microglia prepared from P3 mice had considerably altered gene expression with higher levels of TNFα, CD11b, arginase-1 and VEGF suggesting that culturing may significantly alter microglial properties. In conclusion, age- and sex-specific variances in basal gene expression may enable differential microglial responses to the same stimulus at different ages, perhaps contributing to altered CNS vulnerabilities and/or disease courses.

Microglia play an important role in inflammatory diseases of the central nervous system (CNS). These cells have also been identified in brain neoplasms; however, as of yet their function largely remains unclear. More recent studies designed to characterize further tumor-associated microglia suggest that the immune effector function of these cells may be suppressed in CNS tumors. Furthermore, microglia and macrophages can secrete various cytokines and growth factors that may contribute to the successful immune evasion, growth, and invasion of brain neoplasms. A better understanding of microglia and macrophage function is essential for the development of immune-based treatment strategies against malignant brain tumors.

The nucleotide receptor P2X7 has been shown to modulate LPS-induced macrophage production of numerous inflammatory mediators. Although the C-terminal portion of P2X7 is thought to be essential for multiple receptor functions, little is known regarding the structural motifs that lie within this region. We show here that the P2X7 C-terminal domain contains several apparent protein-protein and protein-lipid interaction motifs with potential importance to macrophage signaling and LPS action. Surprisingly, P2X7 also contains a conserved LPS-binding domain. In this report, we demonstrate that peptides derived from this P2X7 sequence bind LPS in vitro. Moreover, these peptides neutralize the ability of LPS to activate the extracellular signal-regulated kinases (ERK1, ERK2) and to promote the degradation of the inhibitor of κB-α isoform (IκB-α) in RAW 264.7 macrophages. Collectively, these data suggest that the C-terminal domain of P2X7 may directly coordinate several signal transduction events related to macrophage function and LPS action.