Effect of ibogaine on morphine: Induced modifications of palatability

The ability of the potential anti-addictive agent ibogaine to module the morphine-induced modification of quinine and sucrose palatability was assessed utilizing the taste reactivity test. Ibogaine (40mg/kg) was administered 24 hr prior to an injection of morphine (2 mg/kg), followed 30 min later by a 5 min intraoral infusion of 0.05% quinine solution (Experiment 1) or 10% sucrose solution (Experiment 2). Treatment with morphine enhanced the palatability of both quinine and sucrose solution. Morphine reduced the aversiveness of quinine solution during the 5 min of testing and enhanced ingestive responding to sucrose solution, however, only during min 1 of the 5 min test. Pretreatment with ibogaine 24 hr earlier, regardless of treatment condition, also enhanced the palatability of quinine and sucrose solution. However, there was no evidence that ibogaine modulated the effect of morphine on quinine or sucrose palatability

Editorial:LRRK2-Fifteen Years From Cloning to the Clinic

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Parental effects on early life history traits of haddock Melanogrammus aeglefinus

Gametes from five male and three female haddock ( Melanogrammus aeglefinus ) were crossed to produce 15 half-sibling families that were used to evaluate potential parental contributions to early life history variability. Larval morphology at 0 and 5 days post-hatch (dph) and time to starvation in the absence of food were examined. Maternal influences on larval standard length and yolk area were significant at 0 and 5 dph. Paternal effects on larval standard length were significant at 0 and 5 dph, whereas paternal effects on yolk area were only significant at 5 dph. Larval eye diameter was influenced by maternity at day 0 post-hatch and by both maternity and paternity at 5 dph. Myotome height of larvae was subject to maternal and paternal influences at 0 and 5 dph. Growth rate was significantly influenced by both paternity and maternity. Yolk utilization efficiency was significantly influenced by parental interaction, while the time taken for larvae to die in the absence of food was affected only by maternity. Results of this study not only confirm the importance of female contributions to larval development but also indicate a paternal influence on the development and the early life history success of marine fish

The WD40 Domain Is Required for LRRK2 Neurotoxicity

BACKGROUND:Mutations in leucine-rich repeat kinase 2 (LRRK2) are the most common genetic cause of Parkinson disease (PD). LRRK2 contains an "enzymatic core" composed of GTPase and kinase domains that is flanked by leucine-rich repeat (LRR) and WD40 protein-protein interaction domains. While kinase activity and GTP-binding have both been implicated in LRRK2 neurotoxicity, the potential role of other LRRK2 domains has not been as extensively explored. PRINCIPAL FINDINGS:We demonstrate that LRRK2 normally exists in a dimeric complex, and that removing the WD40 domain prevents complex formation and autophosphorylation. Moreover, loss of the WD40 domain completely blocks the neurotoxicity of multiple LRRK2 PD mutations. CONCLUSION:These findings suggest that LRRK2 dimerization and autophosphorylation may be required for the neurotoxicity of LRRK2 PD mutations and highlight a potential role for the WD40 domain in the mechanism of LRRK2-mediated cell death

Insights into the Influence of Specific Splicing Events on the Structural Organization of LRRK2

Leucine-rich repeat kinase 2 (LRRK2) is a large protein of unclear function. Rare mutations in the LRRK2 gene cause familial Parkinson's disease (PD) and inflammatory bowel disease. Genome-wide association studies (GWAS) have revealed significant association of the abovementioned diseases at the LRRK2 locus. Cell and systems biology research has led to potential roles that LRRK2 may have in PD pathogenesis, especially the kinase domain (KIN). Previous human expression studies showed evidence of mRNA expression and splicing patterns that may contribute to our understanding of the function of LRRK2. In this work, we investigate and identified significant regional differences in LRRK2 expression at the mRNA level, including a number of splicing events in the Ras of complex protein (Roc) and C-terminal of Roc domain (COR) of LRRK2, in the substantia nigra (SN) and occipital cortex (OCTX). Our findings indicate that the predominant form of LRRK2 mRNA is full length, with shorter isoforms present at a lower copy number. Our molecular modelling study suggests that splicing events in the ROC/COR domains will have major consequences on the enzymatic function and dimer formation of LRRK2. The implications of these are highly relevant to the broader effort to understand the biology and physiological functions of LRRK2, and to better characterize the role(s) of LRRK2 in the underlying mechanism leading to PD

Doubly Constrained C-terminal of Roc (COR) Domain-Derived Peptides Inhibit Leucine-Rich Repeat Kinase 2 (LRRK2) Dimerization

Missense mutations along the leucine-rich repeat kinase 2 (LRRK2) protein are a major contributor to Parkinson's Disease (PD), the second most commonly occurring neurodegenerative disorder worldwide. We recently reported the development of allosteric constrained peptide inhibitors that target and downregulate LRRK2 activity through disruption of LRRK2 dimerization. In this study, we designed doubly constrained peptides with the objective of inhibiting C-terminal of Roc (COR)-COR mediated dimerization at the LRRK2 dimer interface. We show that the doubly constrained peptides are cell-permeant, bind wild-type and pathogenic LRRK2, inhibit LRRK2 dimerization and kinase activity, and inhibit LRRK2-mediated neuronal apoptosis, and in contrast to ATP-competitive LRRK2 kinase inhibitors, they do not induce the mislocalization of LRRK2 to skein-like structures in cells. This work highlights the significance of COR-mediated dimerization in LRRK2 activity while also highlighting the use of doubly constrained peptides to stabilize discrete secondary structural folds within a peptide sequence.</p

Allosteric Inhibition of Parkinson's-Linked LRRK2 by Constrained Peptides

Leucine-Rich Repeat Kinase 2 (LRRK2) is a large, multidomain protein with dual kinase and GTPase function that is commonly mutated in both familial and idiopathic Parkinson's Disease (PD). While dimerization of LRRK2 is commonly detected in PD models, it remains unclear whether inhibition of dimerization can regulate catalytic activity and pathogenesis. Here, we show constrained peptides that are cell-penetrant, bind LRRK2, and inhibit LRRK2 activation by downregulating dimerization. We further show that inhibited dimerization decreases kinase activity and inhibits ROS production and PD-linked apoptosis in primary cortical neurons. While many ATP-competitive LRRK2 inhibitors induce toxicity and mislocalization of the protein in cells, these constrained peptides were found to not affect LRRK2 localization. The ability of these peptides to inhibit pathogenic LRRK2 kinase activity suggests that disruption of dimerization may serve as a new allosteric strategy to downregulate PD-related signaling pathways.</p

Vitamin B12 modulates Parkinson’s disease LRRK2 kinase activity through allosteric regulation and confers neuroprotection

Missense mutations in Leucine-Rich Repeat Kinase 2 (LRRK2) cause the majority of familial and some sporadic forms of Parkinson’s disease (PD). The hyperactivity of LRRK2 kinase induced by the pathogenic mutations underlies neurotoxicity, promoting the development of LRRK2 kinase inhibitors as therapeutics. Many potent and specific small molecule LRRK2 inhibitors have been reported with promise. However, nearly all inhibitors are ATP competitive – some with unwanted side effects and unclear clinical outcome - alternative types of LRRK2 inhibitors are lacking. Herein we find 5’-deoxyadenosylcobalamin (AdoCbl), a physiological form of the essential micronutrient vitamin B12 as a mixed-type allosteric inhibitor of LRRK2 kinase activity. Multiple assays show that AdoCbl directly binds LRRK2, leading to the alterations of protein conformation and ATP binding in LRRK2. STD-NMR analysis of a LRRK2 homologous kinase reveals the contact sites in AdoCbl that interface with the kinase domain. Furthermore, we provide evidence that AdoCbl modulates LRRK2 activity through disruption of LRRK2 dimerization. Treatment with AdoCbl inhibits LRRK2 kinase activity in cultured cells and brain tissue, and importantly prevents neurotoxicity in primary rodent cultures as well as in transgenic C. elegans and D. melanogaster expressing LRRK2 disease variants. Finally, AdoCbl alleviates deficits in dopamine release sustainability caused by LRRK2 disease variants in mouse models. Our study uncovers vitamin B12 as a novel class of LRRK2 kinase modulator with a distinct mechanism, which can be harnessed to develop new LRRK2-based PD therapeutics in the futur