Parkin-deficient mice are not more sensitive to 6-hydroxydopamine or methamphetamine neurotoxicity

BACKGROUND: Autosomal recessive juvenile parkinsonism (AR-JP) is caused by mutations in the parkin gene which encodes an E3 ubiquitin-protein ligase. Parkin is thought to be critical for protecting dopaminergic neurons from toxic insults by targeting misfolded or oxidatively damaged proteins for proteasomal degradation. Surprisingly, mice with targeted deletions of parkin do not recapitulate robust behavioral or pathological signs of parkinsonism. Since Parkin is thought to protect against neurotoxic insults, we hypothesized that the reason Parkin-deficient mice do not develop parkinsonism is because they are not exposed to appropriate environmental triggers. To test this possibility, we challenged Parkin-deficient mice with neurotoxic regimens of either methamphetamine (METH) or 6-hydroxydopamine (6-OHDA). Because Parkin function has been linked to many of the pathways involved in METH and 6-OHDA toxicity, we predicted that Parkin-deficient mice would be more sensitive to the neurotoxic effects of these agents. RESULTS: We found no signs consistent with oxidative stress, ubiquitin dysfunction, or degeneration of striatal dopamine neuron terminals in aged Parkin-deficient mice. Moreover, results from behavioral, neurochemical, and immunoblot analyses indicate that Parkin-deficient mice are not more sensitive to dopaminergic neurotoxicity following treatment with METH or 6-OHDA. CONCLUSION: Our results suggest that the absence of a robust parkinsonian phenotype in Parkin-deficient mice is not due to the lack of exposure to environmental triggers with mechanisms of action similar to METH or 6-OHDA. Nevertheless, Parkin-deficient mice could be more sensitive to other neurotoxins, such as rotenone or MPTP, which have different mechanisms of action; therefore, identifying conditions that precipitate parkinsonism specifically in Parkin-deficient mice would increase the utility of this model and could provide insight into the mechanism of AR-JP. Alternatively, it remains possible that the absence of parkinsonism in Parkin-deficient mice could reflect fundamental differences between the function of human and mouse Parkin, or the existence of a redundant E3 ubiquitin-protein ligase in mouse that is not found in humans. Therefore, additional studies are necessary to understand why Parkin-deficient mice do not display robust signs of parkinsonism

Altered Anesthetic Sensitivity of Mice Lacking Ndufs4, a Subunit of Mitochondrial Complex I

RDP and AQ were supported by the Howard Hughes Medical Institute (HHMI). AQ was a recipient of MICINN postdoctoral mobility program fellowship from the Spanish Ministerio de Ciencia e Innovación. PGM and MMS were supported by National Institutes of Health (NIH) grant GM58881. These studies were also supported in part by the Mitochondrial Research Guild. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.Anesthetics are in routine use, yet the mechanisms underlying their function are incompletely understood. Studies in vitro demonstrate that both GABAA and NMDA receptors are modulated by anesthetics, but whole animal models have not supported the role of these receptors as sole effectors of general anesthesia. Findings in C. elegans and in children reveal that defects in mitochondrial complex I can cause hypersensitivity to volatile anesthetics. Here, we tested a knockout (KO) mouse with reduced complex I function due to inactivation of the Ndufs4 gene, which encodes one of the subunits of complex I. We tested these KO mice with two volatile and two non-volatile anesthetics. KO and wild-type (WT) mice were anesthetized with isoflurane, halothane, propofol or ketamine at post-natal (PN) days 23 to 27, and tested for loss of response to tail clamp (isoflurane and halothane) or loss of righting reflex (propofol and ketamine). KO mice were 2.5 - to 3- fold more sensitive to isoflurane and halothane than WT mice. KO mice were 2-fold more sensitive to propofol but resistant to ketamine. These changes in anesthetic sensitivity are the largest recorded in a mammal

Selective deletion of cochlear hair cells causes rapid age-dependent changes in spiral ganglion and cochlear nucleus neurons

During nervous system development, critical periods are usually defined as early periods during which manipulations dramatically change neuronal structure or function, whereas the same manipulations in mature animals have little or no effect on the same property. Neurons in the ventral cochlear nucleus (CN) are dependent on excitatory afferent input for survival during a critical period of development. Cochlear removal in young mammals and birds results in rapid death of target neurons in the CN. Cochlear removal in older animals results in little or no neuron death. However, the extent to which hair-cell-specific afferent activity prevents neuronal death in the neonatal brain is unknown. We further explore this phenomenon using a new mouse model that allows temporal control of cochlear hair cell deletion. Hair cells express the human diphtheria toxin (DT) receptor behind the Pou4f3 promoter. Injections of DT resulted in nearly complete loss of organ of Corti hair cells within 1 week of injection regardless of the age of injection. Injection of DT did not influence surrounding supporting cells directly in the sensory epithelium or spiral ganglion neurons (SGNs). Loss of hair cells in neonates resulted in rapid and profound neuronal loss in the ventral CN, but not when hair cells were eliminated at a more mature age. In addition, normal survival of SGNs was dependent on hair cell integrity early in development and less so in mature animals. This defines a previously undocumented critical period for SGN survival

Viral Gene Delivery Selectively Restores Feeding and Prevents Lethality of Dopamine-Deficient Mice

AbstractDopamine-deficient mice (DA−/−), lacking tyrosine hydroxylase (TH) in dopaminergic neurons, become hypoactive and aphagic and die by 4 weeks of age. They are rescued by daily treatment with L-3,4-dihydroxyphenylalanine (L-DOPA); each dose restores dopamine (DA) and feeding for less than 24 hr. Recombinant adeno-associated viruses expressing human TH or GTP cyclohydrolase 1 (GTPCH1) were injected into the striatum of DA−/− mice. Bilateral coinjection of both viruses restored feeding behavior for several months. However, locomotor activity and coordination were partially improved. A virus expressing only TH was less effective, and one expressing GTPCH1 alone was ineffective. TH immunoreactivity and DA were detected in the ventral striatum and adjacent posterior regions of rescued mice, suggesting that these regions mediate a critical DA-dependent aspect of feeding behavior

Parkin-mediated K63-linked polyubiquitination targets misfolded DJ-1 to aggresomes via binding to HDAC6

Sequestration of misfolded proteins into pericentriolar inclusions called aggresomes is a means that cells use to minimize misfolded protein-induced cytotoxicity. However, the molecular mechanism by which misfolded proteins are recruited to aggresomes remains unclear. Mutations in the E3 ligase parkin cause autosomal recessive Parkinson's disease that is devoid of Lewy bodies, which are similar to aggresomes. Here, we report that parkin cooperates with heterodimeric E2 enzyme UbcH13/Uev1a to mediate K63-linked polyubiquitination of misfolded DJ-1. K63-linked polyubiquitination of misfolded DJ-1 serves as a signal for interaction with histone deacetylase 6, an adaptor protein that binds the dynein–dynactin complex. Through this interaction, misfolded DJ-1 is linked to the dynein motor and transported to aggresomes. Furthermore, fibroblasts lacking parkin display deficits in targeting misfolded DJ-1 to aggresomes. Our findings reveal a signaling role for K63-linked polyubiquitination in dynein-mediated transport, identify parkin as a key regulator in the recruitment of misfolded DJ-1 to aggresomes, and have important implications regarding the biogenesis of Lewy bodies

Striatal GPR88 modulates foraging efficiency

The striatum is anatomically and behaviorally implicated in behaviors that promote efficient foraging. To investigate this function, we studied instrumental choice behavior in mice lacking GPR88, a striatum-enriched orphan G-protein-coupled receptor that modulates striatal medium spiny neuron excitability. Our results reveal that hungry mice lacking GPR88 (KO mice) were slow to acquire food-reinforced lever press but could lever press similar to controls on a progressive ratio schedule. Both WT and KO mice discriminated between reward and no-reward levers; however, KO mice failed to discriminate based on relative quantity-reward (1 vs 3 food pellets) or effort (3 vs 9 lever presses). We also demonstrate preference for the high-reward (3 pellet) lever was selectively reestablished when GPR88 expression was restored to the striatum. We propose that GPR88 expression within the striatum is integral to efficient action-selection during foraging.SIGNIFICANCE STATEMENT Evolutionary pressure driving energy homeostasis favored detection and comparison of caloric value. In wild and laboratory settings, neural systems involved in energy homeostasis bias foraging to maximize energy efficiency. This is observed when foraging behaviors are guided by superior nutritional density or minimized caloric expenditure. The striatum is anatomically and functionally well placed to perform the sensory and motor integration necessary for efficient action selection during foraging. However, few studies have examined this behavioral phenomenon or elucidated underlying molecular mechanisms. Both humans and mice with nonfunctional GPR88 have been shown to present striatal dysfunctions and impaired learning. We demonstrate that GPR88 expression is necessary to efficiently integrate effort and energy density information guiding instrumental choice

A new mouse model to study restoration of interleukin-6 (IL-6) expression in a Cre-dependent manner : microglial IL-6 regulation of experimental autoimmune encephalomyelitis

Altres ajuts: "La Marató TV3" 20142210Interleukin-6 (IL-6) is a pleiotropic cytokine that controls numerous physiological processes both in basal and neuroinflammatory conditions, including the inflammatory response to experimental autoimmune encephalomyelitis (EAE). IL-6 is produced by multiple peripheral and central cells, and until now, the putative roles of IL-6 from different cell types have been evaluated through conditional cell-specific IL-6 knockout mice. Nevertheless, these mice probably undergo compensatory responses of IL-6 from other cells, which makes it difficult to assess the role of each source of IL-6. To give some insight into this problem, we have produced a novel mouse model: a conditional reversible IL-6 KO mouse (IL6-DIO-KO). By using double-inverted, open-reading-frame (DIO) technology, we created a mouse line with the loss of Il6 expression in all cells that can be restored by the action of Cre recombinase. Since microglia are one of the most important sources and targets of IL-6 into the central nervous system, we have recovered microglial Il6 expression in IL6-DIO-KO mice through breeding to Cx3cr1 -CreER mice and subsequent injection of tamoxifen (TAM) when mice were 10-16 weeks old. Then, they were immunized with myelin oligodendrocyte glycoprotein 35-55 peptide (MOG) 7 weeks after TAM treatment to induce EAE. Clinical symptoms and demyelination, CD3 infiltration, and gliosis in the spinal cord were evaluated. IL6-DIO-KO mice were resistant to EAE, validating the new model. Restoration of microglial Il6 was sufficient to develop a mild version of EAE-related clinical symptoms and neuropathology. IL6-DIO-KO mouse is an excellent model to understand in detail the role of specific cellular sources of IL-6 within a recovery-of-function paradigm in EAE

An Excitatory Circuit in the Perioculomotor Midbrain for Non-REM Sleep Control.

The perioculomotor (pIII) region of the midbrain was postulated as a sleep-regulating center in the 1890s but largely neglected in subsequent studies. Using activity-dependent labeling and gene expression profiling, we identified pIII neurons that promote non-rapid eye movement (NREM) sleep. Optrode recording showed that pIII glutamatergic neurons expressing calcitonin gene-related peptide alpha (CALCA) are NREM-sleep active; optogenetic and chemogenetic activation/inactivation showed that they strongly promote NREM sleep. Within the pIII region, CALCA neurons form reciprocal connections with another population of glutamatergic neurons that express the peptide cholecystokinin (CCK). Activation of CCK neurons also promoted NREM sleep. Both CALCA and CCK neurons project rostrally to the preoptic hypothalamus, whereas CALCA neurons also project caudally to the posterior ventromedial medulla. Activation of each projection increased NREM sleep. Together, these findings point to the pIII region as an excitatory sleep center where different subsets of glutamatergic neurons promote NREM sleep through both local reciprocal connections and long-range projections

Loss of mitochondrial Ndufs4 in striatal medium spiny neurons mediates progressive motor impairment in a mouse model of leigh syndrome

Altres ajuts: Juan del la Cierva (IJCI-2015-24576)Inability of mitochondria to generate energy leads to severe and often fatal myoencephalopathies. Among these, Leigh syndrome (LS) is one of the most common childhood mitochondrial diseases; it is characterized by hypotonia, failure to thrive, respiratory insufficiency and progressive mental and motor dysfunction, leading to early death. Basal ganglia nuclei, including the striatum, are affected in LS patients. However, neither the identity of the affected cell types in the striatum nor their contribution to the disease has been established. Here, we used a mouse model of LS lacking Ndufs4, a mitochondrial complex I subunit, to confirm that loss of complex I, but not complex II, alters respiration in the striatum. To assess the role of striatal dysfunction in the pathology, we selectively inactivated Ndufs4 in the striatal medium spiny neurons (MSNs), which account for over 95% of striatal neurons. Our results show that lack of Ndufs4 in MSNs causes a non-fatal progressive motor impairment without affecting the cognitive function of mice. Furthermore, no inflammatory responses or neuronal loss were observed up to 6 months of age. Hence, complex I deficiency in MSNs contributes to the motor deficits observed in LS, but not to the neural degeneration, suggesting that other neuronal populations drive the plethora of clinical signs in LS

Loss of Mitochondrial Ndufs4 in Striatal Medium Spiny Neurons Mediates Progressive Motor Impairment in a Mouse Model of Leigh Syndrome

Inability of mitochondria to generate energy leads to severe and often fatal myoencephalopathies. Among these, Leigh syndrome (LS) is one of the most common childhood mitochondrial diseases; it is characterized by hypotonia, failure to thrive, respiratory insufficiency and progressive mental and motor dysfunction, leading to early death. Basal ganglia nuclei, including the striatum, are affected in LS patients. However, neither the identity of the affected cell types in the striatum nor their contribution to the disease has been established. Here, we used a mouse model of LS lacking Ndufs4, a mitochondrial complex I subunit, to confirm that loss of complex I, but not complex II, alters respiration in the striatum. To assess the role of striatal dysfunction in the pathology, we selectively inactivated Ndufs4 in the striatal medium spiny neurons (MSNs), which account for over 95% of striatal neurons. Our results show that lack of Ndufs4 in MSNs causes a non-fatal progressive motor impairment without affecting the cognitive function of mice. Furthermore, no inflammatory responses or neuronal loss were observed up to 6 months of age. Hence, complex I deficiency in MSNs contributes to the motor deficits observed in LS, but not to the neural degeneration, suggesting that other neuronal populations drive the plethora of clinical signs in LS