Hippocampal synaptic failure is an early event in experimental parkinsonism with subtle cognitive deficit.

Learning and memory mainly rely on correct synaptic function in the hippocampus and other brain regions. In Parkinson's disease, subtle cognitive deficits may even precede motor signs early in the disease. Hence, we set out to unravel the earliest hippocampal synaptic alterations associated with human α-synuclein overexpression prior to and soon after the appearance of cognitive deficits in a parkinsonism model. We bilaterally injected adeno-associated viral vectors encoding A53T-mutated human α-synuclein into the substantia nigra of rats, and evaluated them 1, 2, 4 and 16 weeks post-inoculation by immunohistochemistry and immunofluorescence to study degeneration and distribution of α-synuclein in the midbrain and hippocampus. The object location test was used to evaluate hippocampal-dependent memory. Sequential window acquisition of all theoretical mass spectrometry-based proteomics and fluorescence analysis of single-synapse long-term potentiation were used to study alterations to protein composition and plasticity in isolated hippocampal synapses. The effect of L-DOPA and pramipexole on long-term potentiation was also tested. Human α-synuclein was found within dopaminergic and glutamatergic neurons of the ventral tegmental area, and in dopaminergic, glutamatergic and GABAergic axon terminals in the hippocampus from 1 week post-inoculation, concomitant with mild dopaminergic degeneration in the ventral tegmental area. In the hippocampus, differential expression of proteins involved in synaptic vesicle cycling, neurotransmitter release and receptor trafficking, together with impaired long-term potentiation were the first events observed (1 week post-inoculation), preceding cognitive deficits (4 weeks post-inoculation). Later on, at 16 weeks post-inoculation, there was a deregulation of proteins involved in synaptic function, particularly those involved in the regulation of membrane potential, ion balance and receptor signalling. Hippocampal long-term potentiation was impaired before and soon after the onset of cognitive deficits, at 1 and 4 weeks post-inoculation, respectively. L-DOPA recovered hippocampal long-term potentiation more efficiently at 4 weeks post-inoculation than pramipexole, which partially rescued it at both time points. Overall, we found impaired synaptic plasticity and proteome dysregulation at hippocampal terminals to be the first events that contribute to the development of cognitive deficits in experimental parkinsonism. Our results not only point to dopaminergic but also to glutamatergic and GABAergic dysfunction, highlighting the relevance of the three neurotransmitter systems in the ventral tegmental area-hippocampus interaction from the earliest stages of parkinsonism. The proteins identified in the current work may constitute potential biomarkers of early synaptic damage in the hippocampus and hence, therapies targeting these could potentially restore early synaptic malfunction and consequently, cognitive deficits in Parkinson's disease.

Striatal synaptic bioenergetic and autophagic decline in premotor experimental parkinsonism.

Synaptic impairment might precede neuronal degeneration in Parkinson's disease. However, the intimate mechanisms altering synaptic function by the accumulation of presynaptic α-synuclein in striatal dopaminergic terminals before dopaminergic death occurs, have not been elucidated. Our aim is to unravel the sequence of synaptic functional and structural changes preceding symptomatic dopaminergic cell death. As such, we evaluated the temporal sequence of functional and structural changes at striatal synapses before parkinsonian motor features appear in a rat model of progressive dopaminergic death induced by overexpression of the human mutated A53T α-synuclein in the substantia nigra pars compacta, a protein transported to these synapses. Sequential window acquisition of all theoretical mass spectra proteomics identified deregulated proteins involved first in energy metabolism and later, in vesicle cycling and autophagy. After protein deregulation and when α-synuclein accumulated at striatal synapses, alterations to mitochondrial bioenergetics were observed using a Seahorse XF96 analyser. Sustained dysfunctional mitochondrial bioenergetics was followed by a decrease in the number of dopaminergic terminals, morphological and ultrastructural alterations, and an abnormal accumulation of autophagic/endocytic vesicles inside the remaining dopaminergic fibres was evident by electron microscopy. The total mitochondrial population remained unchanged whereas the number of ultrastructurally damaged mitochondria increases as the pathological process evolved. We also observed ultrastructural signs of plasticity within glutamatergic synapses before the expression of motor abnormalities, such as a reduction in axospinous synapses and an increase in perforated postsynaptic densities. Overall, we found that a synaptic energetic failure and accumulation of dysfunctional organelles occur sequentially at the dopaminergic terminals as the earliest events preceding structural changes and cell death. We also identify key proteins involved in these earliest functional abnormalities that may be modulated and serve as therapeutic targets to counterbalance the degeneration of dopaminergic cells to delay or prevent the development of Parkinson's disease.

Cerebral metabolic pattern associated with progressive parkinsonism in non-human primates reveals early cortical hypometabolism.

Dopaminergic denervation in patients with Parkinson's disease is associated with changes in brain metabolism. Cerebral in-vivo mapping of glucose metabolism has been studied in severe stable parkinsonian monkeys, but data on brain metabolic changes in early stages of dopaminergic depletion of this model is lacking. Here, we report cerebral metabolic changes associated with progressive nigrostriatal lesion in the pre-symptomatic and symptomatic stages of the progressive 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) monkey model of Parkinson's Disease. Monkeys (Macaca fascicularis) received MPTP injections biweekly to induce progressive grades of dopamine depletion. Monkeys were sorted according to motor scale assessments in control, asymptomatic, recovered, mild, and severe parkinsonian groups. Dopaminergic depletion in the striatum and cerebral metabolic patterns across groups were studied in vivo by positron emission tomography (PET) using monoaminergic ([11C]-dihydrotetrabenazine; 11C-DTBZ) and metabolic (2-[18F]-fluoro-2-deoxy-d-glucose; 18F-FDG) radiotracers. 11C-DTBZ-PET analysis showed progressive decrease of binding potential values in the striatum of monkeys throughout MPTP administration and the development of parkinsonian signs. 18F-FDG analysis in asymptomatic and recovered animals showed significant hypometabolism in temporal and parietal areas of the cerebral cortex in association with moderate dopaminergic nigrostriatal depletion. Cortical hypometabolism extended to involve a larger area in mild parkinsonian monkeys, which also exhibited hypermetabolism in the globus pallidum pars interna and cerebellum. In severe parkinsonian monkeys, cortical hypometabolism extended further to lateral-frontal cortices and hypermetabolism also ensued in the thalamus and cerebellum. Unbiased histological quantification of neurons in Brodmann's area 7 in the parietal cortex did not reveal neuron loss in parkinsonian monkeys versus controls. Early dopaminergic nigrostriatal depletion is associated with cortical, mainly temporo-parietal hypometabolism unrelated to neuron loss. These findings, together with recent evidence from Parkinson's Disease patients, suggest that early cortical hypometabolism may be associated and driven by subcortical changes that need to be evaluated appropriately. Altogether, these findings could be relevant when potential disease modifying therapies become available.

Galanin (1-15) Enhances the Behavioral Effects of Fluoxetine in the Olfactory Bulbectomy Rat, Suggesting a New Augmentation Strategy in Depression.

BACKGROUND: Selective serotonergic reuptake inhibitors, including fluoxetine (FLX), are the most commonly used for the treatment of major depression. However, they are effective for remission in only 30% of patients. Recently, we observed that Galanin (1-15) [GAL(1-15)] enhanced the antidepressant effects of FLX in naïve animals, suggesting a new augmentation strategy in depression. METHODS: We have analyzed in an animal model of depression, the olfactory bulbectomy (OBX) rats, the effect of GAL(1-15) on FLX-mediated responses in the forced swimming test and the sucrose preference test and the involvement of GAL receptor 2 with its antagonist, M871. We have also studied the corticosterone levels in OBX after the coadministration of GAL(1-15) with FLX. Moreover, we studied whether the effects of GAL(1-15) on FLX actions were mediated via auto- and heteroreceptor 5-HT1A (5-HT1AR), analyzing the binding characteristics, mRNA levels, and functionality of 5-HT1AR in the dorsal hippocampus. RESULTS: GAL(1-15) enhances the antidepressant-like effects induced by FLX in OBX animals in the forced swimming test and the sucrose preference test. The involvement of the GALR2 was demonstrated with M871. Importantly, the mechanism underlying the GAL(1-15)/FLX interactions in the OBX animals involves the 5-HT1AR in the hippocampus at the plasma membrane (increase of affinity and density of 5HT1AR in the DG) and transcriptional (increase of 5HT1AR mRNA levels in DG and CA1) levels. Besides, the coadministration of GAL(1-15) and FLX also reduced OBX-increased corticosterone levels. CONCLUSIONS: The results open the possibility to use GAL(1-15) in combination with FLX as a novel strategy for the treatment of depression.

[18F]-DPA-714 PET as a specific in vivo marker of early microglial activation in a rat model of progressive dopaminergic degeneration.

PURPOSE: To study the feasibility of the in vivo [18F]-DPA-714 TSPO positron emission tomography (PET) to detect glial activation in a rat model of progressive parkinsonism induced by viral-mediated overexpression of A53T mutated human α-synuclein (hα-syn) in the substantia nigra pars compacta (SNpc). METHODS: We conducted a cross-sectional study in a model of progressive parkinsonism. Bilateral intranigral injections with 2/9 adeno-associated viral vectors encoding either hα-syn (AAV-hα-syn) or green fluorescent protein (AAV-GFP) were performed in rats (n = 60). In vivo [18F]-DPA-714 PET imaging was performed at different time points after inoculation (p.i.) of the viral vector (24 and 72 h and 1, 2, 3, and 16 weeks). Images were analyzed to compute values of binding potential (BP) in the SNpc and striatum using a volume of interest (VOI) analysis. Immunohistochemistry of markers of dopaminergic degeneration (tyrosine hydroxylase (TH)), microglia (Iba-1), and astrocytes (GFAP) was carried out. Binding potential (BP) of [18F]-DPA-714 PET in the in vivo PET study was correlated with post-mortem histological markers. RESULTS: In the SNpc of AAV-hα-syn rats, there was higher in vivo [18F]-DPA-714 BP (p < 0.05) and increased number of post-mortem Iba-1+ cells (p < 0.05) from second week p.i. onwards, which were highly correlated (p < 0.05) between each other. These findings antedated the nigral reduction of TH+ cells that occurs since third week p.i. (p < 0.01). In addition, the [18F]-DPA-714 BP was inversely correlated (p < 0.05) with the TH+ cells. In contrast, GFAP+ cells only increased at 16 weeks p.i. and did not correlate with the in vivo results. In the striatum, no changes in the number of Iba-1+ and GFAP+ cells were observed, but an increment in the [18F]-DPA-714 BP was found at 16 weeks p.i. CONCLUSIONS: Our study showed that in vivo PET study with [18F]-DPA-714 is a selective and reliable biomarker of microglial activation and could be used to study preclinical stages of Parkinson's disease (PD) and to monitor the progression of the disease.

Central administration of galanin N-terminal fragment 1-15 decreases the voluntary alcohol intake in rats.

Alcohol consumption is considered a major risk factor for disease and mortality worldwide. In the absence of effective treatments in alcohol use disorders, it is important to find new biological targets that could modulate alcohol consumption. We tested the role of the N-terminal galanin fragment (1-15) [GAL(1-15)] in voluntary ethanol consumption in rats using the two-bottle choice paradigm as well as compare the effects of GAL(1-15) with the whole molecule of GAL. We describe for the first time that GAL(1-15), via central mechanisms, induces a strong reduction in preference and ethanol consumption in rats. These effects were significantly different than GAL. GAL receptor (GALR) 2 was involved in these effects, because the specific GALR2 antagonist M871 blocked GAL(1-15) mediated actions in preference and ethanol intake. Importantly, the mechanism of this action involves changes in GALR expression and also in immediate-early gene C-Fos and receptors-internalization-related gene Rab5 in the striatum. The relevance of the striatum as a target for GAL(1-15) was supported by the effect of GAL(1-15) on the locomotor activity of rats after ethanol administration. These results may give the basis for the development of novel therapeutics strategies using GAL(1-15) analogues for the treatment of alcohol use disorders in humans.

Ratios of proteins in cerebrospinal fluid in Parkinson's disease cognitive decline: prospective study.

BACKGROUND: There is a need for biomarkers of dementia in PD. OBJECTIVES: To determine if the levels of the main CSF proteins and their ratios are associated with deterioration in cognition and progression to dementia in the short to mid term. METHODS: The Parkinson's Progression Markers Initiative database was used as an exploratory cohort, and a center-based cohort was used as a replication cohort. Amyloid ß1-42, total tau, threonine-181 phosphorylated tau, and α-synuclein in the CSF and the ratios of these proteins were assessed. RESULTS: In the Parkinson's Progression Markers Initiative cohort (n = 281), the total tau/amyloid ß1-42, total tau/α-synuclein, total tau/amyloid ß1-42+α-synuclein, and amyloid ß1-42/total tau ratios were associated with a risk of progression to dementia over a 3-year follow-up. In the replication cohort (n = 40), the total tau/α-synuclein and total tau/amyloid ß1-42+α-synuclein ratios were associated with progression to dementia over a 41-month follow-up. CONCLUSION: Ratios of the main proteins found in PD patient brain inclusions that can be measured in the CSF appear to have value as short- to mid-term predictors of dementia. © 2018 International Parkinson and Movement Disorder Society.

Receptor⁻Receptor Interactions in Multiple 5-HT1A Heteroreceptor Complexes in Raphe-Hippocampal 5-HT Transmission and Their Relevance for Depression and Its Treatment.

Due to the binding to a number of proteins to the receptor protomers in receptor heteromers in the brain, the term "heteroreceptor complexes" was introduced. A number of serotonin 5-HT1A heteroreceptor complexes were recently found to be linked to the ascending 5-HT pathways known to have a significant role in depression. The 5-HT1A⁻FGFR1 heteroreceptor complexes were involved in synergistically enhancing neuroplasticity in the hippocampus and in the dorsal raphe 5-HT nerve cells. The 5-HT1A protomer significantly increased FGFR1 protomer signaling in wild-type rats. Disturbances in the 5-HT1A⁻FGFR1 heteroreceptor complexes in the raphe-hippocampal 5-HT system were found in a genetic rat model of depression (Flinders sensitive line (FSL) rats). Deficits in FSL rats were observed in the ability of combined FGFR1 and 5-HT1A agonist cotreatment to produce antidepressant-like effects. It may in part reflect a failure of FGFR1 treatment to uncouple the 5-HT1A postjunctional receptors and autoreceptors from the hippocampal and dorsal raphe GIRK channels, respectively. This may result in maintained inhibition of hippocampal pyramidal nerve cell and dorsal raphe 5-HT nerve cell firing. Also, 5-HT1A⁻5-HT2A isoreceptor complexes were recently demonstrated to exist in the hippocampus and limbic cortex. They may play a role in depression through an ability of 5-HT2A protomer signaling to inhibit the 5-HT1A protomer recognition and signaling. Finally, galanin (1⁻15) was reported to enhance the antidepressant effects of fluoxetine through the putative formation of GalR1⁻GalR2⁻5-HT1A heteroreceptor complexes. Taken together, these novel 5-HT1A receptor complexes offer new targets for treatment of depression.

Tau/α-synuclein ratio and inflammatory proteins in Parkinson's disease: An exploratory study.

BACKGROUND: No CSF or plasma biomarker has been validated for diagnosis or progression of PD. OBJECTIVES: To assess whether the CSF and plasma levels of proteins associated with PD neuropathological inclusions and with neuroinflammation might have value in the diagnosis of PD or in relation to disease severity. METHODS: CSF levels of α-synuclein, amyloid-ß1-42, total tau, and threonine-181 phosphorylated tau, as well as CSF and plasma levels of cytokines (interleukin-1ß, interleukin-2, interleukin, interferon-γ, and tumor necrosis factor α) were studied in 40 PD patients and 40 healthy controls. Plasma levels of cytokines were measured in 51 patients and 26 aditional controls. We also explored the Parkinson's Progression Markers Initiative data set as a replication cohort. RESULTS: CSF levels of α-synuclein, amyloid-ß1-42, and tumor necrosis factor α were lower in patients than in controls, and the total tau/α-synuclein, phosphorylated tau/α-synuclein, total tau/amyloid-ß1-42+α-synuclein, and phosphorylated tau/amyloid-ß1-42+α-synuclein ratios were higher in patients. The best area under the curve value was obtained for the phosphorylated tau/α-synuclein ratio alone (0.86) and also when this was combined with tumor necrosis factor α in CSF (0.91; sensitivity 92.9%, specificity 75% for a cut-off value of ≤ 0.71). Phosphorylated tau/α-synuclein and phosphorylated tau/amyloid-ß1-42+α-synuclein were higher in patients than in controls of the Parkinson's Progression Markers Initiative database. Plasma cytokines did not differ between groups, although interleukin-6 levels were positively correlated with UPDRS-I, -II, and -III scores. CONCLUSIONS: The CSF phosphorylated tau/α-synuclein ratio alone, and in combination with tumor necrosis factor α and plasma interleukin-6 levels, might serve as biomarkers to diagnose PD and monitor its severity. © 2017 International Parkinson and Movement Disorder Society.

Dopamine D4 receptor stimulation prevents nigrostriatal dopamine pathway activation by morphine: relevance for drug addiction.

Morphine is one of the most effective drugs used for pain management, but it is also highly addictive. Morphine elicits acute and long-term adaptive changes at cellular and molecular level in the brain, which play a critical role in the development of tolerance, dependence and addiction. Previous studies indicated that the dopamine D4 receptor (D4 R) activation counteracts morphine-induced adaptive changes of the µ opioid receptor (MOR) signaling in the striosomes of the caudate putamen (CPu), as well as the induction of several Fos family transcription factors. Thus, it has been suggested that D4 R could play an important role avoiding some of the addictive effects of morphine. Here, using different drugs administration paradigms, it is determined that the D4 R agonist PD168,077 prevents morphine-induced activation of the nigrostriatal dopamine pathway and morphological changes of substantia nigra pars compacta (SNc) dopamine neurons, leading to a restoration of dopamine levels and metabolism in the CPu. Results from receptor autoradiography indicate that D4 R activation modulates MOR function in the substantia nigra pars reticulata (SNr) and the striosomes of the CPu, suggesting that these regions are critically involved in the modulation of SNc dopamine neuronal function through a functional D4 R/MOR interaction. In addition, D4 R activation counteracts the rewarding effects of morphine, as well as the development of hyperlocomotion and physical dependence without any effect on its analgesic properties. These results provide a novel role of D4 R agonist as a pharmacological strategy to prevent the adverse effects of morphine in the treatment of pain.

Biomarkers for dementia and mild cognitive impairment in Parkinson's disease.

Cognitive decline is one of the most frequent and disabling nonmotor features of Parkinson's disease. Around 30% of patients with Parkinson's disease experience mild cognitive impairment, a well-established risk factor for the development of dementia. However, mild cognitive impairment in patients with Parkinson's disease is a heterogeneous entity that involves different types and extents of cognitive deficits. Because it is not currently known which type of mild cognitive impairment confers a higher risk of progression to dementia, it would be useful to define biomarkers that could identify these patients to better study disease progression and possible interventions. In this sense, the identification among patients with Parkinson's disease and mild cognitive impairment of biomarkers associated with dementia would allow the early detection of this process. This review summarizes studies from the past 25 years that have assessed the potential biomarkers of dementia and mild cognitive impairment in Parkinson's disease patients. Despite the potential importance, no biomarker has as yet been validated. However, features such as low levels of epidermal and insulin-like growth factors or uric acid in plasma/serum and of Aß in CSF, reduction of cerebral cholinergic innervation and metabolism measured by PET mainly in posterior areas, and hippocampal atrophy in MRI might be indicative of distinct deficits with a distinct risk of dementia in subgroups of patients. Longitudinal studies combining the existing techniques and new approaches are needed to identify patients at higher risk of dementia. © 2016 International Parkinson and Movement Disorder Society.

Monoaminergic PET imaging and histopathological correlation in unilateral and bilateral 6-hydroxydopamine lesioned rat models of Parkinson's disease: a longitudinal in-vivo study.

Carbon-11 labeled dihydrotetrabenazine ((11)C-DTBZ) binds to the vesicular monoamine transporter 2 and has been used to assess nigro-striatal integrity in animal models and patients with Parkinson's disease. Here, we applied (11)C-DTBZ positron emission tomography (PET) to obtain longitudinally in-vivo assessment of striatal dopaminergic loss in the classic unilateral and in a novel bilateral 6-hydroxydopamine (6-OHDA) lesion rat model. Forty-four Sprague-Dawley rats were divided into 3 sub-groups: 1. 6-OHDA-induced unilateral lesion in the medial forebrain bundle, 2. bilateral lesion by injection of 6-OHDA in the third ventricle, and 3. vehicle injection in either site. (11)C-DTBZ PET studies were investigated in the same animals successively at baseline, 1, 3 and 6weeks after lesion using an anatomically standardized volumes-of-interest approach. Additionally, 12 rats had PET and Magnetic Resonance Imaging to construct a new (11)C-DTBZ PET template. Behavior was characterized by rotational, catalepsy and limb-use asymmetry tests and dopaminergic striatal denervation was validated post-mortem by immunostaining of the dopamine transporter (DAT). (11)C-DTBZ PET showed a significant decrease of striatal binding (SB) values one week after the unilateral lesion. At this point, there was a 60% reduction in SB in the affected hemisphere compared with baseline values in 6-OHDA unilaterally lesioned animals. A 46% symmetric reduction over baseline SB values was found in bilaterally lesioned rats at the first week after lesion. SB values remained constant in unilaterally lesioned rats whereas animals with bilateral lesions showed a modest (22%) increase in binding values at the 3rd and 6th weeks post-lesion. The degree of striatal dopaminergic denervation was corroborated histologically by DAT immunostaining. Statistical analysis revealed a high correlation between (11)C-DTBZ PET SB and striatal DAT immunostaining values (r=0.95, p<0.001). The data presented here indicate that (11)C-DTBZ PET may be used to ascertain changes occurring in-vivo throughout the evolution of nigro-striatal dopaminergic neurodegeneration, mainly in the unilateral 6-OHDA lesion rat.

The relationship between atrophy and hypometabolism: is it regionally dependent in dementias?

Neuronal failure leading to dementia in neurodegenerative diseases is evidenced in vivo by functional and structural changes in the brain such as reductions of glucose consumption and volume of grey matter. The earliest phase of cognitive decline and presymptomatic stages of these diseases are heralded by specific patterns of hypometabolism, even in the absence of atrophy, which are currently considered as diagnostic biomarkers. Atrophy is less consistently found as an initial marker of these diseases and is invariably present in moderate to severe stages with a disease-related topography. The relationship between these two markers is not uniform, but in the two diseases in which they have been directly compared, Alzheimer's and Parkinson's disease, altered hypometabolism precedes and exceeds atrophy in most regions. This suggests a two-step degenerative process. In contrast to these findings, the hippocampus skips this pattern and is more structurally than functionally affected, thereby suggesting a different pathological mechanism in this particular area. More studies are needed to disentangle the mechanisms underlying both markers and their relationship in neurodegenerative diseases.

Prevalence of cancer in Parkinson's disease related to R1441G and G2019S mutations in LRRK2.

An inverse relationship between Parkinson's disease (PD) and cancer has been described. However, the association between cancers and genetic forms of PD, in particular the R1441G mutation in the LRRK2 gene, is not well known. The objective of this work was to analyze cancer prevalence in PD patients with R1441G or G2019S mutations in LRRK2, and in idiopathic PD (iPD). A total of 732 patients with PD (70 and 25 carriers of R1441G or G2019S mutations, respectively), and 177 controls, were linked using a population-based cancer registry of the Spanish province of Gipuzkoa. Cancer prevalence was not significantly higher in PD-G2019S carriers (20%) than in PD-R1441G carriers (14.3%), iPD (13.8%), or controls (12.5%). With the exception of a high prevalence of hematological cancers (crude odds ratio of 7.1) in the R1441G group, specific cancer types were not increased in PD mutation carriers. In both the carrier and iPD groups, cancers were diagnosed after the onset of PD. PD patients had a similar prevalence of cancer to control subjects. There was no increased association between G2019S or R1441G mutations and any type of cancer. Although there was a higher prevalence of hematological cancers in the R1441G group, the low number of such cancers overall makes this finding of uncertain significance. There was a slightly higher but not statistically significant prevalence of non-skin cancers in the G2019S group, suggesting that further study to evaluate the association should be undertaken prior to ascribing an increased cancer risk to this population.

Dopamine D4 receptor counteracts morphine-induced changes in µ opioid receptor signaling in the striosomes of the rat caudate putamen.

The mu opioid receptor (MOR) is critical in mediating morphine analgesia. However, prolonged exposure to morphine induces adaptive changes in this receptor leading to the development of tolerance and addiction. In the present work we have studied whether the continuous administration of morphine induces changes in MOR protein levels, its pharmacological profile, and MOR-mediated G-protein activation in the striosomal compartment of the rat CPu, by using immunohistochemistry and receptor and DAMGO-stimulated [35S]GTPγS autoradiography. MOR immunoreactivity, agonist binding density and its coupling to G proteins are up-regulated in the striosomes by continuous morphine treatment in the absence of changes in enkephalin and dynorphin mRNA levels. In addition, co-treatment of morphine with the dopamine D4 receptor (D4R) agonist PD168,077 fully counteracts these adaptive changes in MOR, in spite of the fact that continuous PD168,077 treatment increases the [3H]DAMGO Bmax values to the same degree as seen after continuous morphine treatment. Thus, in spite of the fact that both receptors can be coupled to Gi/0 protein, the present results give support for the existence of antagonistic functional D4R-MOR receptor-receptor interactions in the adaptive changes occurring in MOR of striosomes on continuous administration of morphine.

New molecular mechanisms to explain the neuroprotective effects of insulin-like growth factor II in a cellular model of Parkinson's disease.

INTRODUCTION: One of the hallmarks of Parkinsons Disease (PD) is oxidative distress, leading to mitochondrial dysfunction and neurodegeneration. Insulin-like growth factor II (IGF-II) has been proven to have antioxidant and neuroprotective effects in some neurodegenerative diseases, including PD. Consequently, there isgrowing interest in understanding the different mechanisms involved in the neuroprotective effect of this hormone. OBJECTIVES: To clarify the mechanism of action of IGF-II involved in the protective effect of this hormone. METHODS: The present study was carried out on a cellular model PD based on the incubation of dopaminergic cells (SN4741) in a culture with the toxic 1-methyl-4-phenylpyridinium (MPP+), in the presence of IGF-II. This model undertakes proteomic analyses in order to understand which molecular cell pathways might be involved in the neuroprotective effect of IGF-II. The most important proteins found in the proteomic study were tested by Western blot, colorimetric enzymatic activity assay and immunocytochemistry. Along with the proteomic study, mitochondrial morphology and function were also studied by transmission electron microscopy and oxygen consumption rate. The cell cycle was also analysed using 7AAd/BrdU staining, and flow cytometry. RESULTS: The results obtained indicate that MPP+, MPP++IGF-II treatment and IGF-II, when compared to control, modified the expression of 197, 246 proteins and 207 respectively. Some of these proteins were found to be involved in mitochondrial structure and function, and cell cycle regulation. Including IGF-II in the incubation medium prevents the cell damage induced by MPP+, recovering mitochondrial function and cell cycle dysregulation, and thereby decreasing apoptosis. CONCLUSION: IGF-II improves mitochondrial dynamics by promoting the association of Mitofilin with mitochondria, regaining function and redox homeostasis. It also rebalances the cell cycle, reducing the amount of apoptosis and cell death by the regulation of transcription factors, such as Checkpoint kinase 1.

Early modulation by the dopamine D4 receptor of morphine-induced changes in the opioid peptide systems in the rat caudate putamen.

The peptides dynorphin and enkephalin modulate many physiological processes, such as motor activity and the control of mood and motivation. Their expression in the caudate putamen (CPu) is regulated by dopamine and opioid receptors. The current work was designed to explore the early effects of the acute activation of D4 and/or µ opioid receptors by the agonists PD168,077 and morphine, respectively, on the regulation of the expression of these opioid peptides in the rat CPu, on transcription factors linked to them, and on the expression of µ opioid receptors. In situ hybridization experiments showed that acute treatment with morphine (10 mg/kg) decreased both enkephalin and dynorphin mRNA levels in the CPu after 30 min, but PD168,077 (1 mg/kg) did not modify their expression. Coadministration of the two agonists demonstrated that PD168,077 counteracted the morphine-induced changes and even increased enkephalin mRNA levels. The immunohistochemistry studies showed that morphine administration also increased striatal µ opioid receptor immunoreactivity but reduced P-CREB expression, effects that were blocked by the PD168,077-induced activation of D4 receptors. The current results present evidence of functional D4 -µ opioid receptor interactions, with consequences for the opioid peptide mRNA levels in the rat CPu, contributing to the integration of DA and opioid peptide signaling.

l-dopa-induced dyskinesias in unilateral 6-hydroxydopamine-lesioned rats are not modified by excitotoxic lesion of the entopeduncular nucleus and substantia nigra pars reticulata.

l-Dopa-induced dyskinesias (LIDs) are a troublesome complication in Parkinson's disease after long-term therapy and a major reason for surgical treatment. LIDs are effectively eliminated by surgery. We aimed to reproduce such effect in the 6-hydroxydopamine (6-OHDA)-lesioned rat model. Single or combined lesions with quinolinic acid were caused in the entopeduncular nucleus (EP) and substantia nigra pars reticulata (SNr) on 6-hydroxydopamine (6-OHDA)-lesioned rats treated for 3 weeks with l-Dopa (6 mg/kg plus 15 mg/kg benserazide, i.p.). l-Dopa administration was continued for a further week following the lesion and abnormal involuntary movements (AIMs) scored at the end of treatment. Neither the individual lesions of the EP and SNr nor the combined lesions had any antidyskinetic effect nor decreased the total number of rotations. These results suggest that excitotoxic lesions of neurons bodies of the output nuclei of the basal ganglia, which destroy cell bodies and spare fibers of passage, do not induce a beneficial reduction of dyskinesias in contrast to thermolytic lesions in humans (which provokes a complete tissue destruction), thus supporting the possibility that other nuclei or systems might be involved in the antidyskinetic effect of pallidotomy.

Teaching the physiology of the human body in non-formal spaces: pilot experience of a Service-Learning methodology and the interaction between students of different educational levels.

Research institutes and universities have strengthened the development of biomedicine outreach activities, constituing a non-formal education system of science literacy, although with little commitment from undergraduate students. However, as a Service-Learning methodology, these outreach activities could work as a tool for the acquisition of skills by students of Health Science Degrees. Described here is the development of the workshop entitled "Exploring the human body" at the Biodonostia Health Research Institute and the pilot experience of its implementation as a Service-Learning activity at the University of Málaga. Firstly, 359 secondary education students were mentored by Ph.D. students through a 5-station workshop with experiments and activities related to the physiology of the human body. Then, 301 undergraduate students of Medicine and Nursing Degrees advised 965 secondary education students. Both groups of students assessed the workshop via questionnaires and a debriefing. The data showed an overall score of 4.6 out of 5 for the workshop. Undergraduate students reported a positive impact on their academic background (4.8 out of 5), mainly due to the improvement of oral communication skills (34%). Therefore, this methodology could be a valid and applicable tool to develop the cross-disciplinary competences of undergraduate students.

The effects of reward and frustration in patients with bipolar disorder: Evidence from a computerized task with non-contingent feedback.

BACKGROUND: Bipolar disorder (BD) is characterized by mood changes that implies alterations in reward sensitivity and frustration tolerance. This study examined the effects of monetary reward and frustration on attentional performance and on affective experience across mood states in BD. METHODS: An Affective Posner Task in which the nature of contingencies are divided in the three successive blocks (baseline condition, monetary reward and non-contingent feedback) was applied to BD individuals in their different episodes: mania (n = 30), depression (n = 30), and euthymia (n =  30) as well as to a group of healthy controls (n = 30). RESULTS: Monetary reward improved performance (in terms of faster response times) in the euthymic group and the control group, whereas it impaired performance in the manic group and has not significant effect in the depressed group. In addition, an increased interference of frustration on response accuracy was exhibited in the three groups of BD patients (including euthymia) compared with healthy controls. LIMITATIONS: Participants' affective experience was self-informed by a Likert scale, so the reliability of this measure can be undermined in symptomatic patients in terms of stability and objectivity. Although it was statistically controlled, at the time of testing, all BD patients were medicated. CONCLUSIONS: A dissociated effect of reward and frustration was found between symptomatic and euthymic states in BD: whereas the benefit from monetary reward is affected only during symptomatic episodes (i.e., a state), the notably increased interference of frustration is exhibited also during euthymia (i.e., a trait).