Acute inflammation causes morphological and functional rearrangements of mitochondia in astrocytes

Inflammation is thought to contribute to the pathogenesis of neurodegenerative diseases. Among the resident population of cells in the brain, astroglia have been suggested to actively participate in the induction and regulation of neuroinflammation by controlling the secretion of local mediators. However, the initial cellular mechanisms by which astrocytes react to pro-inflammatory molecules are still unclear. our study identified mitochondia as high sensitive organelles that rapidly respond to inflammatory stimuly. Time-lapse video microscopy revealed that mitochondrial morphology, dynamics and motility are drastically altered upon inflammation, resulting in perinuclear clustering of mitochondria. These mitochondrial rearrangements are accompanied by an increased formation of reactive oxygen species and a recruitment of autophagic vacuoles. 24 to 48 hours after the acute inflammatory stimulus, however, the mitochondrial network is re-established. Strikingly, the recovery of a tubular mitochondrial network is abolished in astrocytes with a defective autophagic response, indicating that activation of autophagy is required to restore mitochondrial dynamics. By employing co-cultivation assays we observed that primary cortical neurons undergo degeneration in the presence of inflamed astrocytes. However, this effect was not observed when the primary neurons were grown in conditioned medium derived from inflamed astrocytes, suggesting that a direct contact between astrocytes and neurons mediates neuronal dysfunction upon inflammation. Our results suggest that astrocytes react to inflammatory stimuli by transiently rearranging their mitochondria, a process that involves the autophagic machinery

Inflammatory stimuli prompt autophagic events in cortical astrocytes.

An impaired inflammatory response represents the initial starting point for many neurodegenerative diseases. For their capability of releasing and being target as well of pro-inflammatory molecules and growth factors, astrocytes can be considered key players participating in the development of these neu-rodegenerative processes. Moreover, astroglial cells cover unique functions in the adult brain parenchyma by regulating synaptic transmission and, therefore, contributing to the maintenance of neuronal homeostasis. To investigate how astrocytes react to the initial phases of inflammation, we cultured rat cortical astrocytes and exposed them to pro-inflammatory molecules such as lipopolysaccharide (LPS) and interferon-gamma (IFN-gamma). We observed that stimulated cells rapidly (8-24h) acquire typical features of reactive atrocytes, by increasing the expression of markers such as glial fibrillary acidic pro-tein (GFAP) and inducible nitric oxide synthase (iNOS) evaluated by immunocytochemistry and im-munoblotting, respectively. Surprisingly, LPS/IFN-gamma treatment did not influence cell viability, meas-ured by MTT test, even at later time-points, but inflammation altered astrocyte morphology inducing the formation of vacuoles characteristic of autophagic processes. We analyzed the expression of the au-tophagic marker Microtubule Associated Protein 1 Light Chain (LC3B) over time and correlated it with an altered morphology of mitochondria in these cells by time-lapse microscopy. Our data demonstrate that astrocytes rapidly respond to inflammation by undergoing autophagy. Further experiments will reveal the molecular pathways involved in this reaction and what role these events have in controlling the inflammatory response

Pink1-deficiency in mice impairs gait, olfaction and serotonergic innervation of the olfactory bulb

Parkinson's Disease (PD) is the most common neurodegenerative movement disorder. Autosomal-recessive mutations in the mitochondrial protein kinase PINK1 (PTEN-induced kinase 1) account for 1-2% of the hereditary early-onset cases. To study the mechanisms underlying disease development, we generated Pink1-deficient mice. In analogy to other genetic loss-of-function mouse models, Pink1(-/-) mice did not show morphological alterations in the dopaminergic system. As a consequence, no gross motor dysfunctions were observed indicating that these mice do not develop the cardinal symptoms of PD. Nonetheless, symptoms which develop mainly before bradykinesia, rigidity and resting tremor were clearly evident in Pink1-deficient mice. These symptoms were gait alterations and olfactory dysfunctions. Remarkably in the glomerular layer of the olfactory bulb the density of serotonergic fibers was significantly reduced. Concerning mitochondrial morphology, neurons in Pink1(-/-) mice had less fragmented mitochondria. In contrast, upon acute knock-down of Pink1 increased mitochondrial fragmentation was observed in neuronal cultures. This fragmentation was, however, evened out within days. Taken together, we demonstrate that Pink1-deficient mice exhibit behavioral symptoms of early phases of PD and present systematic experimental evidence for compensation of Pink1-deficiency at the cellular level. Thus, Pink1-deficient mice represent a model for the early phases of PD in which compensation may still impede the onset of neurodegeneration. Consequently, these mice are a valuable tool for studying Pink1-related PD development, as well as for searching for reliable PD biomarkers

The E3 ligase Parkin maintains mitochondrial integrity by increasing linear ubiquitination of NEMO.

Parkin, a RING-between-RING-type E3 ubiquitin ligase associated with Parkinson's disease, has a wide neuroprotective activity, preventing cell death in various stress paradigms. We identified a stress-protective pathway regulated by parkin that links NF-kappa B signaling and mitochondrial integrity via linear ubiquitination. Under cellular stress, parkin is recruited to the linear ubiquitin assembly complex and increases linear ubiquitination of NF-kappa B essential modulator (NEMO), which is essential for canonical NF-kappa B signaling. As a result, the mitochondrial guanosine triphosphatase OPA1 is transcriptionally upregulated via NF-kappa B-responsive promoter elements for maintenance of mitochondrial integrity and protection from stress-induced cell death. Parkin-induced stress protection is lost in the absence of either NEMO or OPA1, but not in cells defective for the mitophagy pathway. Notably, in parkin-deficient cells linear ubiquitination of NEMO, activation of NF-kappa B, and upregulation of OPA1 are significantly reduced in response to TNF-alpha stimulation, supporting the physiological relevance of parkin in regulating this antiapoptotic pathway

The E3 Ligase Parkin Maintains Mitochondrial Integrity by Increasing Linear Ubiquitination of NEMO

Parkin, a RING-between-RING-type E3 ubiquitin ligase associated with Parkinson's disease, has a wide neuroprotective activity, preventing cell death in various stress paradigms. We identified a stress-protective pathway regulated by parkin that links NF-kB signaling and mitochondrial integrity via linear ubiquitination. Under cellular stress, parkin is recruited to the linear ubiquitin assembly complex and increases linear ubiquitination of NF-kB essential modulator (NEMO), which is essential for canonical NF-kB signaling. As a result, the mitochondrial guanosine triphosphatase OPA1 is transcriptionally upregulated via NF-kB-responsive promoter elements for maintenance of mitochondrial integrity and protection from stress-induced cell death. Parkin-induced stress protection is lost in the absence of either NEMO or OPA1, but not in cells defective for the mitophagy pathway. Notably, in parkin-deficient cells linear ubiquitination of NEMO, activation of NF-kB, and upregulation of OPA1 are significantly reduced in response to TNF-alpha stimulation, supporting the physiological relevance of parkin in regulating this antiapoptotic pathway