Based on the results from genetic, biochemical and pharmacological studies presented here, we propose a working model () by which LPS and TNF stimulate microglia and macrophages by binding TLR4 and TNF receptors, respectively to result in activation of NF-κB-dependent gene transcription in activated cells. Activation of this important pathway transiently decreases Parkin mRNA and protein levels in microglia and macrophages, suggesting that dynamic regulation of Parkin levels may subserve an important role of Parkin in these cell types that may include limiting expression of genes involved in inflammatory responses, as suggested by our QPCR analyses. Microglia and macrophages respond to external stimuli in order to perform their immune surveillance role in the central nervous system (CNS) and peripheral circulation, respectively and they activate the NF-κB signaling pathway in response to LPS and TNF treatment 
. Although NF-κB activates the expression of many target genes, it has also been reported to repress gene transcription, including that of the rat androgen receptor 
. Moreover, the Drosophila melanogaster
homolog of NF-κB/Rel (dorsal
), positively regulates some genes while negatively regulating others during embryonic development 
is reported to act as a repressor when it associates with a neighboring co-repressor. The LPS-induced repression of Parkin expression may occur through a similar mechanism involving a co-repressor. The identification of a putative NF-κB site in the mouse parkin
promoter is a novel observation, and to our knowledge this is the first demonstration that Parkin transcription is repressed by inflammatory stimuli in microglia, macrophages and neurally differentiated dopaminergic neuroblastoma cells.
Proposed model of LPS-induced and NF-κB-dependent Parkin downregulation.
Importantly, our finding that LPS and TNF-dependent NF-κB signaling, which serves to activate microglia, also represses Parkin levels suggests a complex interaction between neuroinflammatory signaling and Parkin. One possible result of this interaction is that a transient decrease in Parkin levels may be necessary for normal activation and proliferation of microglia and macrophages following LPS or TNF stimulation. In support of this idea, Parkin expression was found to be transcriptionally repressed in dividing cells 
. Although this instance of repression was mediated by N-myc, this example demonstrates a cellular mechanism of Parkin downregulation during cellular proliferation. A similar example of regulation through repression occurs in apoptotic signaling cascades when specific members of the inhibitor of apoptosis protein (IAP) family are sequestered or destroyed in order for apoptotic signaling to proceed 
. While it is possible that dynamic changes in Parkin levels subserve an important and as yet undefined role of Parkin in microglial activation, complete loss of Parkin may contribute to dysregulated microglial responses in the CNS. In support of this idea, our data that LPS-treated Parkin-null macrophages displayed heightened inflammation-related gene expression suggests potentially important functional consequences as a result of complete loss of Parkin in monocytes. Conversely, microglia or macrophages with atypically high levels of Parkin (such as those from TNF-null mice) displayed blunted activation responses, in agreement with studies that show macrophages from TNF-null mice display weakened activation responses when challenged with oxidative stress 
. Lastly, Parkin-null mice have been reported to display increased microglial proliferation and activation in the CNS 
and mitochondrial alterations in glial cells have been associated with an inability to provide trophic support to neuronal cultures 
The functional consequences of acute and chronic downregulation of Parkin levels in microglia and macrophages and its impact on DA neurons will be addressed in future studies; however, several important predictions can be made on the basis of our findings and our model. O-glycosylated α-synuclein (αSp22) has been shown to be a Parkin substrate 
and one expected consequence of chronic downregulation of Parkin levels is likely to be accumulation of Parkin substrates. Consistent with this prediction is the observation that the rate of clearance of α-synuclein aggregates in BV2 microglia is significantly reduced after treatment of the cells with LPS 
. Although the mechanism by which LPS attenuated α-synuclein clearance in activated microglia in those studies was not explored, our findings suggest that downregulation of Parkin levels by LPS-induced NF-κB signaling might in part account for the accumulation of α-synuclein and perhaps other Parkin substrates; this possibility will need to be tested directly. Another important prediction based on our in vitro
findings is that chronic neuroinflammation and persistent activation of NF-κB in the CNS during CNS infection has the potential to elicit a sustained reduction in Parkin levels, which could also lead to dysregulation of microglia and a toxic microenvironment for DA neurons. Alternatively, repeated exposure to environmental toxins and/or the aging process itself, both of which are associated with chronic neuroinflammation in the CNS, may also result in sustained reductions of Parkin levels in the CNS. All of these environmental triggers could essentially phenocopy the effect of parkin
loss, thereby increasing the vulnerability to peripheral inflammation-induced nigral dopaminergic pathway degeneration 
and may predispose an individual to development of PD.
Given that loss of dopamine (DA)-producing neurons in the substantia nigra pars compacta is the neuropathological hallmark of PD, it is understandable that research in the field has focused on analyzing neuronal phenotypes in PD patients and models of PD-like pathology. However, recent studies suggest the exciting possibilities that genes that influence the risk for development of PD may have important roles in glial cells, and are involved in innate immune function that determine how brain immune surveillance cells respond to insult. Nurr1, a transcription factor involved in the development and maintenance of DA neurons 
, was recently reported to be a critical regulator of cytokine expression in microglia and astrocytes. Elegant studies demonstrated that reduction in Nurr1 expression resulted in excess cytokine production (including TNF) that led to death of DA neurons in LPS-treated mice 
. These unexpected findings underscore two important ideas: the glial microenvironment surrounding DA neurons is a critical determinant of neuronal health and survival; and functions of ubiquitously expressed genes initially thought to be important only in neurons may also have important roles in glial cells and are likely to be dynamically regulated. In addition, genes involved in histocompatibility and immunocompetence have been identified as risk loci for PD in recent genome-wide association studies (GWAS) 
. Taken together, these exciting findings provide compelling rationale to seek a deeper understanding of how inflammatory signaling and innate immune processes influence the functional outcome of neuron-glia interactions and contribute to or protect against development of PD, the second most common neurodegenerative disease in the U.S.