Enhancement of Haloperidol-Induced Catalepsy by GPR143, an L-Dopa Receptor, in Striatal Cholinergic Interneurons.

Dopamine neurons play crucial roles in pleasure, reward, memory, learning, and fine motor skills and their dysfunction is associated with various neuropsychiatric diseases. Dopamine receptors are the main target of treatment for neurologic and psychiatric disorders. Antipsychotics that antagonize the dopamine D2 receptor (DRD2) are used to alleviate the symptoms of these disorders but may also sometimes cause disabling side effects such as parkinsonism (catalepsy in rodents). Here we show that GPR143, a G-protein-coupled receptor for L-3,4-dihydroxyphenylalanine (L-DOPA), expressed in striatal cholinergic interneurons enhances the DRD2-mediated side effects of haloperidol, an antipsychotic agent. Haloperidol-induced catalepsy was attenuated in male Gpr143 gene-deficient (Gpr143-/y ) mice compared with wild-type (Wt) mice. Reducing the endogenous release of L-DOPA and preventing interactions between GPR143 and DRD2 suppressed the haloperidol-induced catalepsy in Wt mice but not Gpr143-/y mice. The phenotypic defect in Gpr143-/y mice was mimicked in cholinergic interneuron-specific Gpr143-/y (Chat-cre;Gpr143flox/y ) mice. Administration of haloperidol increased the phosphorylation of ribosomal protein S6 at Ser240/244 in the dorsolateral striatum of Wt mice but not Chat-cre;Gpr143flox/y mice. In Chinese hamster ovary cells stably expressing DRD2, co-expression of GPR143 increased cell surface expression level of DRD2, and L-DOPA application further enhanced the DRD2 surface expression. Shorter pauses in cholinergic interneuron firing activity were observed after intrastriatal stimulation in striatal slice preparations from Chat-cre;Gpr143flox/y mice compared with those from Wt mice. Together, these findings provide evidence that GPR143 regulates DRD2 function in cholinergic interneurons and may be involved in parkinsonism induced by antipsychotic drugs.

L-DOPA Receptor GPR143 Functionally Couples with Adrenergic α1B Receptor at the Second Transmembrane Interface.

Adrenergic receptors (ADRs) are widely distributed in the peripheral and central nervous systems. We previously reported that L-3,4-dihydroxyphenylalanine (L-DOPA), the precursor of dopamine, sensitizes adrenergic α1 receptor (ADRA1) through a G protein-coupled receptor GPR143. Chimeric analysis, in which the transmembrane (TM) domains of GPR143 were replaced with those of GPR37, revealed that the second TM region was essential for the potentiation of phenylephrine-induced extracellular signal-regulated kinase (ERK) phosphorylation by GPR143. In HEK293T cells expressing ADRA1B, phenylephrine-induced ERK phosphorylation was augmented by the co-expression of GPR143, compared to the mock vector. Immunoprecipitation analysis revealed that a synthetic transactivator of the transcription peptide fused with TM2 of GPR143 (TAT-TM2) disrupts the interaction between GPR143 and ADRA1B. This TAT-TM2 peptide suppressed the augmentation of phenylephrine-induced ERK phosphorylation by GPR143 in HEK293T cells co-expressing ADRA1B and GPR143. These results indicate that the interaction between GPR143 and ADRA1B is required for the potentiation of ADRA1B-mediated signaling by GPR143. The TM2 region of GPR143 is a crucial dimeric interface for the functional coupling between ADRA1B and GPR143.

Involvement of the L-DOPA receptor GPR143 in acute and chronic actions of methylphenidate.

Methylphenidate (MPH) and methamphetamine (METH) are the current treatments of choice for attention deficit/hyperactivity disorder. We previously reported that METH induces the release of dopamine (DA) and of the neurotransmitter candidate L-3,4-dihydroxyphenylalanine (L-DOPA). In contrast, we here found that MPH increased the DA release while it did not affect the L-DOPA release from the dorsolateral striatum. Nevertheless, MPH-induced hyperlocomotion was reduced in Gpr143 (L-DOPA receptor) gene-deficient (Gpr143-/y) mice. The rewarding effect and increased c-fos expression induced by MPH were also attenuated in Gpr143-/y mice. Together, these findings suggest that GPR143 is involved in the acute and chronic actions of MPH.

Coupling between GPR143 and dopamine D2 receptor is required for selective potentiation of dopamine D2 receptor function by L-3,4-dihydroxyphenylalanine in the dorsal striatum.

Dopamine (DA) is involved in neurological and physiological functions such as motor control. L-3,4-dihydroxyphenylalanine (L-DOPA), a precursor of DA, is conventionally believed to be an inert amino acid precursor of DA, and its major therapeutic effects in Parkinson's disease (PD) are mediated through its conversion to DA. On the contrary, accumulating evidence suggests that L-DOPA itself is a neurotransmitter. We here show that L-DOPA potentiates DA D2 receptor (DRD2) signaling through GPR143, the gene product of X-linked ocular albinism 1, a G-protein-coupled receptor for L-DOPA. In Gpr143-gene-deficient (Gpr143-/y ) mice, quinpirole, a DRD2/DRD3 agonist, -induced hypolocomotion was attenuated compared to wild-type (WT) mice. Administration of non-effective dose of L-DOPA methyl ester augmented the quinpirole-induced hypolocomotion in WT mice but not in Gpr143-/y mice. In cells co-expressing GPR143 and DRD2, L-DOPA enhanced the interaction between GPR143 and DRD2 and augmented quinpirole-induced decrease in cAMP levels. This augmentation by L-DOPA was not observed in cells co-expressing GPR143 and DRD1 or DRD3. Chimeric analysis in which the domain of GPR143 was replaced with GPR37 revealed that GPR143 interacted with DRD2 at the fifth transmembrane domain. Intracerebroventricular administration of a peptide that disrupted the interaction mitigated quinpirole-induced behavioral changes in WT mice but not in Gpr143-/y mice. These findings provide evidence that coupling between GPR143 and DRD2 is required for selective DRD2 modulation by L-DOPA in the dorsal striatum.

L-DOPA-Induced Neurogenesis in the Hippocampus Is Mediated Through GPR143, a Distinct Mechanism of Dopamine.

Neurogenesis occurs in the hippocampus throughout life and is implicated in various physiological brain functions such as memory encoding and mood regulation. L-3,4-dihydroxyphenylalanine (L-DOPA) has long been believed to be an inert precursor of dopamine. Here, we show that L-DOPA and its receptor, GPR143, the gene product of ocular albinism 1, regulate neurogenesis in the dentate gyrus (DG) in a dopamine-independent manner. L-DOPA at concentrations far lower than that of dopamine promoted proliferation of neural stem and progenitor cells in wild-type mice under the inhibition of its conversion to dopamine; this effect was abolished in GPR143 gene-deficient (Gpr143-/y) mice. Hippocampal neurogenesis decreased during development and adulthood, and exacerbated depression-like behavior was observed in adult Gpr143-/y mice. Replenishment of GPR143 in the DG attenuated the impaired neurogenesis and depression-like behavior. Our findings suggest that L-DOPA through GPR143 modulates hippocampal neurogenesis, thereby playing a role in mood regulation in the hippocampus.

Right ventricular overloading is attenuated in monocrotaline-induced pulmonary hypertension model rats with a disrupted Gpr143 gene, the gene that encodes the 3,4-l-dihydroxyphenyalanine (l-DOPA) receptor.

Pulmonary hypertension (PH) is a severe and progressive disease that causes elevated right ventricular systolic pressure, right ventricular hypertrophy and ultimately right heart failure. However, the underlying pathophysiologic mechanisms are poorly understood. We previously showed that 3,4-l-dihydroxylphenyalanine (DOPA) sensitizes vasomotor response to sympathetic tone via coupling between the adrenergic receptor alpha1 (ADRA1) and a G protein-coupled receptor 143 (GPR143), a DOPA receptor. We investigated whether DOPA similarly enhances ADRA1-mediated contraction in pulmonary arteries isolated from rats, and whether GPR143 is involved in the PH pathogenesis. Pretreating the isolated pulmonary arteries with DOPA 1 µM enhanced vasoconstriction in response to phenylephrine, an ADRA1 agonist, but not to U-46619, a thromboxane A2 agonist or endothelin-1. We generated Gpr143 gene-deficient (Gpr143-/y) rats, and confirmed that DOPA did not augment phenylephrine-induced contractile response in Gpr143-/y rat pulmonary arteries. We utilized a rat model of monocrotaline (MCT)-induced PH. In the MCT model, the right ventricular systolic pressure was attenuated in the Gpr143-/y rats than in WT rats. Phenylephrine-induced cell migration and proliferation were also suppressed in Gpr143-/y pulmonary artery smooth muscle cells than in WT cells. Our result suggests that GPR143 is involved in the PH pathogenesis in the rat models of PH.

Involvement of A5/A7 noradrenergic neurons and B2 serotonergic neurons in nociceptive processing: a fiber photometry study.

In the central nervous system, the A6 noradrenaline (NA) and the B3 serotonin (5-HT) cell groups are well-recognized players in the descending antinociceptive system, while other NA/5-HT cell groups are not well characterized. A5/A7 NA and B2 5-HT cells project to the spinal horn and form descending pathways. We recorded G-CaMP6 green fluorescence signal intensities in the A5/A7 NA and the B2 5-HT cell groups of awake mice in response to acute tail pinch stimuli, acute heat stimuli, and in the context of a non-noxious control test, using fiber photometry with a calcium imaging system. We first introduced G-CaMP6 in the A5/A7 NA or B2 5-HT neuronal soma, using transgenic mice carrying the tetracycline-controlled transactivator transgene under the control of either a dopamine ß-hydroxylase or a tryptophan hydroxylase-2 promoters and by the site-specific injection of adeno-associated virus (AAV-TetO(3G)-G-CaMP6). After confirming the specific expression patterns of G-CaMP6, we recorded G-CaMP6 green fluorescence signals in these sites in awake mice in response to acute nociceptive stimuli. G-CaMP6 fluorescence intensity in the A5, A7, and B2 cell groups was rapidly increased in response to acute nociceptive stimuli and soon after, it returned to baseline fluorescence intensity. This was not observed in the non-noxious control test. The results indicate that acute nociceptive stimuli rapidly increase the activities of A5/A7 NA or B2 5-HT neurons but the non-noxious stimuli do not. The present study suggests that A5/A7 NA or B2 5-HT neurons play important roles in nociceptive processing in the central nervous system. We suggest that A5/A7/B2 neurons may be new therapeutic targets. All performed procedures were approved by the Institutional Animal Use Committee of Kagoshima University (MD17105) on February 22, 2018.

Dual orexin receptor antagonist (DORA-12) treatment affects the overall levels of Net/maoA mRNA expression in the hippocampus.

The orexinergic system plays a significant role in regulating proper sleep/wake maintenance. Dual orexin receptor antagonist (DORA) is widely prescribed for insomnia symptoms. The antagonist acts on orexin 1 and 2 receptors located in certain brain areas, including the locus coeruleus and dorsal raphe. Nevertheless, its effects on monoamine-related gene expression remain unclear. Here, we measured the expression levels of monoamine-related genes in DORA-treated mice. DORA treatment significantly affected overall levels of noradrenalin transporter/monoamine oxidases A mRNA expression in the hippocampus. Our findings suggest that DORA contributes to noradrenalin-related gene expression regulation in the central nervous system.

Distribution of mRNA for GPR143, a receptor of 3,4-L-dihydroxyphenylalanine, and of immunoreactivities for nicotinic acetylcholine receptors in the nigrostriatal and mesolimbic regions.

Nicotine exerts its reinforcing actions by activating nicotinic acetylcholine receptors (nAChRs), but the detailed mechanisms remain unclear. Nicotine releases 3, 4-dihydroxyphenylalanine (DOPA), a neurotransmitter candidate in the central nervous system. Here, we investigated the distribution of GPR143, a receptor of DOPA, and nAChR subunits in the nigrostriatal and mesolimbic regions. We found GPR143 mRNA-positive cells in the striatum and nucleus accumbens. Some of them were surrounded by tyrosine hydroxylase (TH)-immunoreactive fibers. There were some GPR143 mRNA-positive cells coexpressing TH, and nAChR subunit α4 or α7 in the substantia nigra and ventral tegmental area. These findings suggest that DOPA-GPR143 signaling may be involved in the nicotine action in the nigrostriatal and mesolimbic dopaminergic systems.

Enhancement of the rewarding effects of 3,4-methylenedioxymethamphetamine in orexin knockout mice.

Orexinergic neurons, which are closely associated with narcolepsy, regulate arousal and reward circuits through the activation of monoaminergic neurons. Psychostimulants as well as 5-HT-related compounds have potential in the treatment of human narcolepsy. Previous studies have demonstrated that orexin receptor antagonists as well as orexin deficiencies affect the pharmacological effects of psychostimulants. However, little information is available on the consequences of psychostimulant use under orexin deficiency. Therefore, the present study was designed to investigate the abuse liability of psychostimulants in orexin knockout (KO) mice. In the present study, conditioned place preferences induced by methamphetamine and methylphenidate were not altered in orexin KO mice. Interestingly, we found that MDMA induced a conditioned place preference in orexin KO mice, but not in wild type (WT) mice. In addition, MDMA produced methylphenidate/methamphetamine-like discriminative stimulus effects in orexin KO mice, but not WT mice. Increases in 5-HT and dopamine release in the nucleus accumbens induced by MDMA were not altered by knockout of orexin; the steady-state level of G protein activation was higher in the limbic forebrain of orexin KO mice. In substitution tests using a drug discrimination procedure, substitution of 5-HT1A receptor agonist for the discriminative stimulus effects of methylphenidate was enhanced in orexin KO mice. These findings indicate that the orexinergic system is involved the rewarding effects of psychostimulants. However, there is a risk of establishing rewarding effects of psychostimulants even under orexin deficiency. On the other hand, deficiencies in orexin may enhance the abuse liability of MDMA by changing a postsynaptic signal transduction accompanied by changes in discriminative stimulus effects themselves.

Genetic associations of single nucleotide polymorphisms in the l-DOPA receptor (GPR143) gene with severity of nicotine dependence in Japanese individuals, and attenuation of nicotine reinforcement in Gpr143 gene-deficient mice.

l-3,4-dihydroxyphenylalanine (l-DOPA) is a candidate neurotransmitter. l-DOPA is released by nicotine through nicotinic receptors. Recently, G-protein coupled receptor GPR143, was identified as a receptor for l-DOPA. In this study, genetic association studies between GPR143 genetic polymorphisms and smoking behaviors revealed that the single-nucleotide polymorphism rs6640499, in the GPR143 gene, was associated with traits of smoking behaviors in Japanese individuals. In Gpr143 gene-deficient mice, nicotine-induced hypolocomotion and rewarding effect were attenuated compared to those in wild-type mice. Our findings suggest the involvement of GPR143 in the smoking behaviors.

Ziprasidone, a second-generation antipsychotic, affects core clock gene mRNA expression in mice.

Some psychiatric diseases are associated with disruptions in the circadian clock system. Ziprasidone (ZIP), a second-generation antipsychotic, is widely used for psychiatry-related pharmacotherapy but its mechanism has not been clearly elucidated. We measured clock gene fluctuation patterns in the hippocampus and the amygdala in ZIP-treated mice. ZIP significantly increased Per1, Per2, and Bmal1 mRNA 2 h after the lights were turned off (ZT14) in the hippocampus, but not in the amygdala. These results suggest that ZIP might affect clock gene regulation, which could represent the pathway underlying symptom amelioration.

Involvement of A13 dopaminergic neurons located in the zona incerta in nociceptive processing: a fiber photometry study.

The roles of serotonergic and noradrenergic signaling in nociceptive processing in the central nervous system are well known. However, dopaminergic signaling is also relevant to various physical functions, including nociception. The zona incerta is a subthalamic nucleus in which the A13 dopaminergic cell group resides, but how this A13 group affects nociceptive processing remains unknown. Recently, we showed that acute nociceptive stimuli rapidly induce the activity of A10 (ventral tegmental area) dopamine neurons via fiber photometry. In this study, we measured the activity of A13 dopaminergic neurons in response to acute nociceptive stimuli using the same system. Adeno-associated viruses (AAV-CAG-FLEX-G-CaMP6 and AAV-CAG-FLEX-mCherry) were unilaterally injected into the A13 site in transgenic mice carrying a dopamine transporter promotor-regulated Cre recombinase transgene to specifically introduce G-CaMP6/mCherry into A13 dopaminergic cell bodies through site-specific infection. We measured G-CaMP6/mCherry fluorescence intensity in the A13 site to acute nociceptive stimuli (pinch stimulus and heat stimulus). These stimuli significantly induced a rapid increase in G-CaMP6 fluorescence intensity, but non-nociceptive control stimuli did not. In contrast, mCherry fluorescence intensity was not significantly changed by nociceptive stimuli or non-nociceptive stimuli. Our finding is the first report to measure the activity of A13 dopaminergic neurons to aversive stimuli. A13 dopaminergic neurons project to the periaqueductal gray and the central nucleus of the amygdala, which are both well known as key regions in nociceptive processing. Therefore, together with our A10 study, our results indicate that A13 dopaminergic neurons play important roles in nociceptive processing.

Lanthionine ketimine ester improves outcome in an MPTP-induced mouse model of Parkinson's disease via suppressions of CRMP2 phosphorylation and microglial activation.

Parkinson's disease (PD) is characterized by progressive degeneration of dopaminergic neurons in the substantia nigra pars compacta (SNc). Levodopa (L-Dopa), the current main treatment for PD, reduces PD symptoms by partially replacing dopamine, but it does not slow neurodegeneration. Recent studies have evidenced that neuroinflammatory processes contribute to the degeneration of dopaminergic neurons in the SNc under cytopathic conditions, while other lines of inquiry have implicated phosphorylation of collapsin response mediator protein 2 (CRMP2) as a causal factor in axonal retraction after neural injury. We recently reported on the therapeutic effect of lanthionine ketimine ester (LKE) which associates with CRMP2 following axonal injury in the spinal cord. In the present study, we report that LKE protects SNc dopaminergic neurons after 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) challenge, a common model for PD, and reduces the number of activated microglia proximal to the damaged SNc. The results also show that MPTP-induced motor impairment was suppressed in LKE treatment. Furthermore, the results show that LKE inhibits the elevation of CRMP2 phosphorylation in dopaminergic neurons in the SNc after MPTP injection. These data suggest that modification of CRMP2 phosphorylation and suppression of microglial activation with LKE administration may represent a novel strategy for slowing progress of pathological processes in PD.

Low Incidence of High-Grade Pancreatic Intraepithelial Neoplasia Lesions in a Crmp4 Gene-Deficient Mouse Model of Pancreatic Cancer.

Pancreatic intraepithelial neoplasia (PanIN), the most common premalignant lesion of the pancreas, is a histologically well-defined precursor to invasive pancreatic ductal adenocarcinoma (PDAC). However, the molecular mechanisms underlying the progression of PanINs have not been fully elucidated. Previously, we demonstrated that the expression of collapsin response mediator protein 4 (CRMP4) in PDAC was associated with poor prognosis. The expression of CRMP4 was also augmented in a pancreatitis mouse model. However, the role of CRMP4 in the progression of PanIN lesions remains uncertain. In the present study, we examined the relationship between CRMP4 expression and progression of PanIN lesions using genetically engineered mouse models. PanIN lesions were induced by peritoneal injection of the cholecystokinin analog caerulein in LSL-KRASG12D; Pdx1-Cre (KC-Crmp4 wild-type, WT) mice and LSL-KRASG12D; Pdx1-Cre; Crmp4-/- (KC-Crmp4 knockout, KO) mice. We analyzed pancreatic tissue sections from these mice and evaluated PanIN grade by hematoxylin and eosin staining. CRMP4 expression was examined and the cellular components assessed by immunohistochemistry using antibodies against CRMP4, CD3, and α-smooth muscle actin (SMA). The incidence of high-grade PanIN in KC-Crmp4 WT mice was higher than that in KC-Crmp4 KO animals. CRMP4 was expressed not only in epithelial cells but also in αSMA-positive cells in stromal areas of PanIN lesions. The CRMP4 expression in stromal areas correlated with PanIN grade in WT mice. These results suggested that the expression of CRMP4 in stromal cells may underlie the incidence or progression of PanIN.

Involvement of supralemniscal nucleus (B9) 5-HT neuronal system in nociceptive processing: a fiber photometry study.

Nociception is important perception that has harmful influence on daily life of humans. As to main pain management system, some descending pathways are called descending antinociceptive systems (DAS). As main pathways of DAS, it is well known that dorsal raphe (B6/B7) - rostral ventromedial medulla (B3) - spinal dorsal horn includes serotonergic system. However, possible role of supralemniscal (B9) serotonin (5-HT) cell group in pain management is still open question. In this study, we measured activities of B9 5-HT neuronal cell bodies and B9 5-HT neuron-derived axons located in the locus coeruleus (LC) and ventral tegmental area (VTA), which are also main players of pain management, using fiber photometry system. We introduced the G-CaMP6 in B9 5-HT neurons using transgenic mice carrying a tetracycline-controlled transactivator transgene (tTA) under the control of a tryptophan hydroxylase-2 (TPH2) promoter and site-specific injection of adeno associated virus (AAV-TetO(3G)-G-CaMP6). After confirmation of specific expression of G-CaMP6 in the target population, G-CaMP6 fluorescence intensity in B9 group and LC/VTA groups was measured in awake mice exposed to acute tail pinch and heat stimuli. G-CaMP6 fluorescence intensity rapidly increased by both stimuli in all groups, but not significantly reacted by nonnociceptive control stimuli. The present results clearly indicate that acute nociceptive stimuli cause a rapid increase in the activities of B9-LC/B9-VTA 5-HTergic pathways, suggesting that B9 5-HT neurons play important roles in nociceptive processing.

Overexpression of the gene product of ocular albinism 1 (GPR143/OA1) but not its mutant forms inhibits neurite outgrowth in PC12 cells.

Neurite outgrowth is a complex differentiation process regulated by external and/or internal mechanisms. Among external mechanisms, G-protein coupled receptors (GPCRs) have been implicated in this process, but the pathways involved are not fully understood. L-3,4-dihydroxyphenylalanine (l-DOPA) is considered to be inert by itself, and to relieve Parkinson's disease through its conversion to dopamine. We have proposed that l-DOPA acts as a neurotransmitter. GPR143, the gene product of ocular albinism 1 (OA1), was identified as a receptor for l-DOPA. OA1 is an X-linked disorder characterized by all typical visual anomalies associated with hypopigmentation and optic misrouting, resulting in severe reduction of visual acuity. However, the molecular basis for this phenotype remains unknown. To study the function of GPR143, we investigated the phenotypic effect of overexpression of GPR143 in pheochromocytoma (PC12) cells treated with nerve growth factor. Overexpression of mouse GPR143 inhibited neurite outgrowth, and the effect was mitigated by l-DOPA cyclohexylester, an antagonist for l-DOPA. Furthermore, knockdown of G-protein Gα13 attenuated mouse GPR143 induced inhibition of neurite outgrowth. Human wild-type (wt) GPR143 also inhibited neurite outgrowth, but its mutants did not mimic the effect of wt GPR143. Our results provide a mechanism for axon guidance phenotype in ocular albinism 1.

l-DOPA and Its Receptor GPR143: Implications for Pathogenesis and Therapy in Parkinson's Disease.

l-3,4-Dihydroxyphenylalanine (l-DOPA) is the most effective therapeutic agent for Parkinson's disease (PD). l-DOPA is traditionally believed to be an inert amino acid that exerts actions and effectiveness in PD through its conversion to dopamine. In contrast to this generally accepted idea, l-DOPA is proposed to be a neurotransmitter. Recently, GPR143 (OA1), the gene product of ocular albinism 1 was identified as a receptor candidate for l-DOPA. GPR143 is widely expressed in the central and peripheral nervous system. GPR143 immunoreactivity was colocalized with phosphorylated α-synuclein in Lewy bodies in PD brains. GPR143 may contribute to the therapeutic effectiveness of l-DOPA and might be related to pathogenesis of PD.

Network-guided analysis of hippocampal proteome identifies novel proteins that colocalize with Aß in a mice model of early-stage Alzheimer's disease.

Alzheimer's disease (AD) is an incurable neurodegenerative disease characterized by memory loss and neurotoxic amyloid beta (Aß) plaques accumulation. Numerous pharmacological interventions targeting Aß plaques accumulation have failed to alleviate AD. Also, the pathological alterations in AD start years before the onset of clinical symptoms. To identify proteins at play during the early stage of AD, we conducted proteomic analysis of the hippocampus of young AppNL-F mice model of AD at the preclinical phase of the disease. This was followed by interactome ranking of the proteome into hubs that were further validated in vivo using immunoblot analysis. We also performed double-immunolabeling of these hub proteins and Aß to quantify colocalization. Behavioral analysis revealed no significant difference in memory performance between 8-month-old AppNL-F and control mice. The upregulation and downregulation of several proteins were observed in the AppNL-F mice compared to control. These proteins corresponded to pathways and processes related to Aß clearance, inflammatory-immune response, transport, mitochondrial metabolism, and glial cell proliferation. Interactome analysis revealed several proteins including DLGP5, DDX49, CCDC85A, ADCY6, HEPACAM, HCN3, PPT1 and TNPO1 as essential proteins in the AppNL-F interactome. Validation by immunoblot confirmed the over-expression of these proteins except HCN3 in the early-stage AD mice hippocampus. Immunolabeling revealed a significant increase in ADCY6/Aß and HEPACAM/Aß colocalized puncta in AppNL-F mice compared to WT. These data suggest that these proteins may be involved in the early stage of AD. Our work suggests new targets and biomarkers for AD diagnosis and therapeutic intervention.

Genetic suppression of collapsin response mediator protein 2 phosphorylation improves outcome in methyl-4-phenyl-1,2,3,6-tetrahydropyridine-induced Parkinson's model mice.

Parkinson's disease (PD) is a common neurodegenerative disorder characterized by slow and progressive degeneration of dopaminergic neurons in the substantia nigra pars compacta (SNc). Levodopa (l-Dopa), the current main treatment for PD, supplies dopamine, but it does not prevent neurodegeneration. There is thus no promising remedy for PD. Recent in vitro study showed the increase in the phosphorylation levels of Collapsin Response Mediator Protein 2 (CRMP2) is involved in dopaminergic axon degeneration. In the present study, we report elevation of CRMP2 phosphorylation in dopaminergic neurons in SNc after challenge with the dopaminergic neurotoxin 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP), a common model for PD. Genetic suppression of CRMP2 phosphorylation by mutation of the obligatory Cyclin-dependent kinase 5 (Cdk5)-targeted serine-522 site prevented axonal degradation in the nigrostriatal pathway of transgenic mice. As a result, the degree of MPTP-induced motor impairment in the rotarod test was suppressed. These results suggest that suppression of CRMP2 phosphorylation may be a novel therapeutic target for PD.