Salivary ATP13A2 is a potential marker of therapy-induced motor complications and is expressed by inclusions in submandibulary glands in Parkinson ´s disease.

Background: ATP13A2 holds promise as biomarker for Parkinsons disease (PD). No study has examined how salivary ATP13A2 is related to motor features in idiopathic PD. Methods: Salivary ATP13A2 concentration was evaluated with ELISA, and statistical correlations of ATP13A2 level with PD parameters were examined. The dose intensity of the dopaminergic medication regimen was expressed as levodopa equivalent daily dose (LEDD). ATP13A2 expression on histological sections of submandibular glands was evaluated using immunohistochemistry. Results: Salivary ATP13A2 was undetectable in many subjects (28 % of patients, 43.7 % of controls). However, all the patients with motor complications (n = 28) showed quantifiable levels of ATP13A2, that positively correlated with MDS-UPDRS (total, parts III and IV), and LEDD (p < 0.05). Dyskinetic patients showed the highest LEDD values (p < 0.05). The histological study revealed: a) ATP13A2 staining was very intense in duct cells and vascular endothelium, and b) two patterns of ATP13A2-positive deposits are observed: rounded inclusions of 10-20 µm in diameter located in the interlobular tissue of the patients, and amorphous aggregates inside duct lumen in controls and patients. Conclusions: The sensitivity of the ELISA assay was a major limitation for quantifying ATP13A2. However, salivary ATP13A2 was detected in all patients with motor complications, where a direct relationship among ATP13A2 concentration, levodopa equivalent daily dose, and MDS-UPDRS was found. Therefore, salivary ATP13A2 might be a reliable index of therapy-induced motor complications. ATP13A2 was expressed by rounded inclusions in the submandibulary gland of patients. This is the first description of ATP13A2-positive inclusions outside the nervous system.

Myeloperoxidase and Advanced Oxidation Protein Products in the Cerebrospinal Fluid in Women and Men with Parkinson's Disease.

Background: Myeloperoxidase (MPO) and advanced oxidation protein products, or AOPP (a type of MPO-derived chlorinated adducts), have been implicated in Parkinson´s disease (PD). Human MPO also show sex-based differences in PD. The objective was to study the relationship of MPO and AOPP in the cerebrospinal fluid (CSF) with motor features of idiopathic PD in male and female patients. Methods: MPO concentration and activity and AOPP content were measured in the CSF and serum in 34 patients and 30 controls. CSF leukocytes and the integrity of the blood-brain barrier were evaluated. Correlations of MPO and AOPP with clinical variables were examined. Results: The blood-brain barrier was intact and CSF leukocyte count was normal in all patients. CSF MPO concentration and activity were similar in the cohort of patients and controls, but CSF MPO content was significantly higher in male patients than in PD women (p = 0.0084). CSF MPO concentration correlated with disease duration in male and female patients (p < 0.01). CSF MPO concentration was significantly higher in men with disease duration ≥12 years versus the remainder of the male subjects (p < 0.01). Changes in CSF MPO in women were not significant. Serum MPO concentration and activity were significantly higher in all PD patients relative to controls (p < 0.0001). CSF MPO was not correlated with serum MPO. Serum AOPP were detected in all patients, but CSF AOPP was undetectable in 53% of patients. AOPP were not quantifiable in controls. Conclusions: CSF MPO is not a good biomarker for PD because mean CSF MPO concentration and activity are not different between the cohort of patients and controls. CSF MPO concentration positively correlated with disease duration in men and women, but CSF MPO is significantly enhanced only in male patients with disease duration longer than 12 years. It can be hypothesized that the MPO-related immune response in early-stage PD might be weak in all patients, but then the MPO-related immune response is progressively enhanced in men, not women. Since the blood-brain barrier is intact, and CSF MPO is not correlated with serum MPO, CSF myeloperoxidase would reflect MPO content in brain cells, not blood-derived cells. Finally, serum AOPP was detected in all patients, but not controls, which is consistent with the occurrence of chlorinative stress in blood serum in PD. The study of CSF AOPP as biomarker could not be assessed because the ELISA assay was hampered by its detection limit in the CSF.

ATP13A2 levels in serum and cerebrospinal fluid in patients with idiopathic Parkinson's disease.

BACKGROUND: The enzyme ATP13A2 holds promise as biomarker in Parkinson's disease (PD). No study has examined the content of ATP13A2 in serum and cerebrospinal fluid (CSF) in idiopathic PD cohorts, or how ATP13A2 relates to the clinical features of the disease. METHODS: ATP13A2 concentration was evaluated with ELISA and immunoblotting. Correlations of serum and CSF ATP13A2 with clinical parameters were examined. The antiparkinsonian medication regimen was expressed as levodopa equivalent dose (LED, mg/day). RESULTS: Serum ATP13A2 concentration was similar in patients and controls, and it correlated with LED and MDS-UPDRS part-IV score (p < .0001), a scale which allows evaluating motor complications. LED also correlated with MDS-UPDRS part-IV score (p < .0001). Serum ATP13A2 concentration and LED were higher in patients with motor complications than in patients without motor complications (p < .0001). The ratio of serum ATP13A2 concentration versus LED was calculated, and mean value was similar in patients with or without motor complications. ATP13A2 concentration in the CSF was undetectable in many subjects because the ELISA assay was hampered by its detection limit. Immunoblotting indicated that CSF ATP13A2 content was higher in patients relative to controls (p = .0002), and no clinical correlations were found. CONCLUSIONS: Increasing LED enhanced serum ATP13A2 concentration and facilitated the development of motor complications. There is a direct relationship between serum ATP13A2 level and the dose intensity of the antiparkinsonian dopaminergic medication. The associations between serum ATP13A2 and LED suggest that serum ATP13A2 content might be a marker of dopamine replacement therapy.

Native α-Synuclein, 3-Nitrotyrosine Proteins, and Patterns of Nitro-α-Synuclein-Immunoreactive Inclusions in Saliva and Submandibulary Gland in Parkinson's Disease.

Background. Salivary α-synuclein (aSyn) and its nitrated form, or 3-nitrotyrosine-α-synuclein (3-NT-αSyn), hold promise as biomarkers for idiopathic Parkinson's disease (IPD). Nitrative stress that is characterized by an excess of 3-nitrotyrosine proteins (3-NT-proteins) has been proposed as a pathogenic mechanism in IPD. The objective is to study the pathological role of native αSyn, 3-NT-αSyn, and 3-NT-proteins in the saliva and submandibulary glands of patients with IPD. Methods. The salivary and serum αSyn and 3-NT-proteins concentration is evaluated with ELISA in patients and controls. Correlations of αSyn and 3-NT-proteins content with clinical features of the disease are examined. Immunohistochemical 3-NT-αSyn expression in submandibulary gland sections is analyzed. Results. (a) Salivary concentration and saliva/serum ratios of native αSyn and 3-NT-proteins are similar in patients and controls; (b) salivary αSyn and 3-NT-proteins do not correlate with any clinical feature; and (c) three patterns of 3-NT-αSyn-positive inclusions are observed on histological sections: rounded "Lewy-type" aggregates of 10-25 µm in diameter, coarse deposits with varied morphology, and spheroid inclusions or bodies of 3-5 µm in diameter. "Lewy-type" and coarse inclusions are observed in the interlobular connective tissue of the gland, and small-sized bodies are located within the cytoplasm of duct cells. "Lewy-type" inclusions are only observed in patients, and the remaining patterns of inclusions are observed in both the patients and controls. Conclusions. The patients' saliva presents a similar concentration of native αSyn and 3-nitrotyrosine-proteins than that of the controls, and no correlations with clinical features are found. These findings preclude the utility of native αSyn in the saliva as a biomarker, and they indicate the absence of nitrative stress in the saliva and serum of patients. As regards nitrated αSyn, "Lewy-type" inclusions expressing 3-NT-αSyn are observed in the patients, not the controls-a novel finding that suggests that a biopsy of the submandibulary gland, if proven safe, could be a useful technique for diagnosing IPD. Finally, to our knowledge, this is also the first description of 3-NT-αSyn-immunoreactive intracytoplasmic bodies in cells that are located outside the nervous system. These intracytoplasmic bodies are present in duct cells of submandibulary gland sections from all subjects regardless of their pathology, and they can represent an aging or involutional change. Further immunostaining studies with different antibodies and larger samples are needed to validate the data.

A Cross-Sectional Study of Foot Growth and Its Correlation with Anthropometric Parameters in a Representative Cohort of Schoolchildren from Southern Spain.

Background: The relationship between growth of the foot and other anthropometric parameters during body development until puberty has been scarcely studied. Some studies propose that growth of the foot in length may be an early index of puberty. The objective of this cross-sectional study was to analyze the relationship between the growth of the foot in length and width with other anthropometric parameters, in prepubertal and early pubertal schoolchildren (Tanner stage II). Methods: Using an instrument that was designed and calibrated for this purpose, maximum foot length, width and height were obtained in 1005 schoolchildren. Results: The findings indicate that the age of onset of pubertal foot growth spur was 7-8 years in girls, and 8-9 years in boys. Growth in foot length stabilized in both sexes after 12 years of age. In boys, a strong correlation was found between height and foot length (r = 0.884; p < 0.047), and between body mass index (BMI) and forefoot width at 12 years of age (r = 0.935; p < 0.020). A strong correlation was found between height and forefoot width at 6 years in girls (r = 0.719; p < 0.001), as well as between BMI and metatarsal width in 10 years-old girls (r = 0.812; p <0.001). Conclusions: The average increase in foot length and width that precedes the onset of Tanner's stage II in both girls and boys can be considered as a useful biological indicator of the onset of puberty.

Cerebrospinal fluid lactoperoxidase level is enhanced in idiopathic Parkinson's disease, and correlates with levodopa equivalent daily dose.

Lactoperoxidase (LPO) is proposed to play a role in the pathogenesis of Parkinson's disease (PD). This enzyme has been reported to be enhanced in the cerebrospinal fluid (CSF) in parkinsonian patients. The objective was to look at the relationship of LPO in the CSF and serum with clinical features of idiopathic PD. LPO concentration was analyzed through ELISA techniques. Correlation of CSF or serum LPO and MDS-UPDRS, dopaminergic medication, and other clinical parameters was examined. The findings revealed that LPO concentration in the CSF, not serum, was found to be elevated in patients with PD relative to controls (p < 0.001). CSF LPO concentration negatively correlated with MDS-UPDRS part-IV score (p < .0001), a rating scale that allows evaluating motor complications. CSF LPO level inversely correlated with the dose intensity of the dopaminergic medication regimen, as evaluated with levodopa equivalent dose or LED (mg/day; p < .0001). LED value positively correlated with MDS-UPDRS part-IV score (p < .0001). To sum up, the findings indicate that CSF LPO is found to be elevated in the CSF of PD patients, and this enzyme holds promise as potential biomarker for diagnosis of PD. Increasing the dose intensity of the dopaminergic medication regimen attenuates the elevation in LPO levels in the CSF, and it facilitates the development of motor complications in patients. The pathophysiological mechanisms that seem to be responsible for LPO increase would include dopamine deficiency, oxidative stress, and less likely, microbial infection.

A Rare Paronychia with Superinfection with Prevotella bivia and Staphylococcus haemolyticus: The Importance of Early Microbiological Diagnosis.

Prevotella bivia is an anaerobic, gram-negative bacillus which naturally thrives in the human vagina, and is usually related to vaginal tract infections. However, this microorganism can also cause infections in other body locations. Infections with Prevotella bivia are frequently severe due to the risk of osteomyelitis and the lack of good protocols for adequate therapeutic management. Staphylococcus haemolyticus infection is one of the most frequent etiological factors of nosocomial infections, which hasthe ability to acquire multiple resistance against antimicrobial agents. We report a rare case of foot and hand paronychia with superinfection of Prevotella bivia and Staphylococcus haemolyticus. We highlight the importance of early microbiological diagnosis, and proper therapeutic management to avoid the risk of complications and the development of bacterial resistance to antibiotics.

Oleoylethanolamide and Palmitoylethanolamide Protect Cultured Cortical Neurons Against Hypoxia.

Introduction: Perinatal hypoxic-ischemic (HI) encephalopathy is defined as a neurological syndrome where the newborn suffers from acute ischemia and hypoxia during the perinatal period. New therapies are needed. The acylethanolamides, oleoylethanolamide (OEA) and palmitoylethanolamide (PEA), possess neuroprotective properties, and they could be effective against perinatal HI. These lipid mediators act through peroxisome proliferator-activated receptors subtype α (PPARα), or transient receptor potential vanilloid (TRPV), such as TRPV subtype 1 and 4. Materials and Methods: The objectives of this study were to discern: (1) the neuroprotective role of OEA and PEA in parietotemporal cortical neurons of newborn rats and mice subjected to hypoxia, and (2) the role of the receptors, PPARα, TRPV1, and TRPV4, in neuroprotective effects. Cell culture of cortical neurons and the lactate dehydrogenase assay was carried out. The role of receptors was discerned by using selective antagonist and agonist ligands, as well as knockout (KO) PPARα mice. Results: The findings indicate that OEA and PEA exert neuroprotective effects on cultured cortical neurons subjected to a hypoxic episode. These protective effects are not mediated by the receptors, PPARα, TRPV1, or TRPV4, because neither PPARα KO mice nor receptor ligands significantly modify OEA and PEA-induced effects. Blocking TRPV4 with RN1734 is neuroprotective per se, and cotreatment with OEA and PEA is able to enhance neuroprotective effects of the acylethanolamides. Since stimulating TRPV4 was devoid of effects on OEA and PEA-induced protective effects, effects of RN1734 cotreatment seem to be a consequence of additive actions. Conclusion: The lipid mediators, OEA and PEA, exert neuroprotective effects on cultured cortical neurons subjected to hypoxia. Coadministration of OEA or PEA, and the TRPV4 antagonist RN1734 is able to enhance neuroprotective effects. These in vitro results could be of utility for developing new therapeutic tools against perinatal HI.

Palmitoylethanolamide prevents neuroinflammation, reduces astrogliosis and preserves recognition and spatial memory following induction of neonatal anoxia-ischemia.

RATIONAL: Neonatal anoxia-ischemia (AI) particularly affects the central nervous system. Despite the many treatments that have been tested, none of them has proven to be completely successful. Palmitoylethanolamide (PEA) and oleoylethanolamide (OEA) are acylethanolamides that do not bind to CB1 or CB2 receptors and thus they do not present cannabinoid activity. These molecules are agonist compounds of peroxisome proliferator-activator receptor alpha (PPARα), which modulates the expression of different genes that are related to glucose and lipid metabolism, inflammation, differentiation and proliferation. OBJECTIVE: In the present study, we analyzed the effects that the administration of PEA or OEA, after a neonatal AI event, has over different areas of the hippocampus. METHODS: To this end, 7-day-old rats were subjected to AI and then treated with vehicle, OEA (2 or 10 mg/kg) or PEA (2 or 10 mg/kg). At 30 days of age, animals were subjected to behavioral tests followed by immunohistochemical studies. RESULTS: Results showed that neonatal AI was associated with decreased locomotion, as well as recognition and spatial memory impairments. Furthermore, these deficits were accompanied with enhanced neuroinflammation and astrogliosis, as well as a decreased PPARα expression. PEA treatment was able to prevent neuroinflammation, reduce astrogliosis and preserve cognitive functions. CONCLUSIONS: These results indicate that the acylethanolamide PEA may play an important role in the mechanisms underlying neonatal AI, and it could be a good candidate for further studies regarding neonatal AI treatments.

Excess amounts of 3-iodo-l-tyrosine induce Parkinson-like features in experimental approaches of Parkinsonism.

3-iodo-l-tyrosine might play a role in Parkinson's disease since this molecule is able, at high concentration, to inhibit tyrosine-hydroxylase activity, the rate-limiting enzyme in dopamine biosynthesis. The possible Parkinson-like effects of 3-iodo-l-tyrosine were tested on three experimental approaches in mice: cultured substantia nigra neurons, the enteric nervous system of the jejunum after intra-peritoneal infusions, and the nigrostriatal system following unilateral intrabrain injections. 3-iodo-l-tyrosine, a physiological molecule, was used at concentrations higher than its serum levels in humans. Parkinson-like signs were evaluated through abnormal aggregation of α-synuclein and tyrosine-hydroxylase, loss of tyrosine-hydroxylase-expressing and striatum-projecting neurons and fibers, reduced tyrosine-hydroxylase density, and Parkinson-like motor and non-motor deficits. The retrograde tracer FluoroGold was used in the brain model. The findings revealed that excess amounts of 3-iodo-l-tyrosine induce Parkinson-like effects in the three experimental approaches. Thus, culture neurons of substantia nigra show, after 3-iodo-l-tyrosine exposure, intracytoplasmic inclusions that express α-synuclein and tyrosine-hydroxylase. Intra-peritoneal infusions of 3-iodo-l-tyrosine cause, in the long-term, α-synuclein aggregation, thicker α-synuclein-positive fibers, and loss of tyrosine-hydroxylase-positive cells and fibers in intramural plexuses and ganglia of the jejunum. Infusion of 3-iodo-l-tyrosine into the left dorsal striata of mice damages the nigrostriatal system, as revealed through lower striatal tyrosine-hydroxylase density, reduced number of tyrosine-hydroxylase-expressing and striatum-projecting neurons in the left substantia nigra, as well as the emergence of Parkinson-like behavioral deficits such as akinesia, bradykinesia, motor disbalance, and locomotion directional bias. In conclusion, excess amounts of 3-iodo-l-tyrosine induce Parkinson-like features in cellular, enteric and brain approaches of Parkinsonism in mice.

Analysis of neurotrophic and antioxidant factors related to midbrain dopamine neuronal loss and brain inflammation in the cerebrospinal fluid of the elderly.

Midbrain dopamine neuronal loss and neuroinflammation are two phenomena that are associated with brain senescence. Neurotrophic factor changes and oxidative stress could subserve these phenomena. Aging-related brain changes can be well monitored through the cerebrospinal fluid (CSF). The objective was to analyze neurotrophic and oxidative parameters that could be related to midbrain dopamine neuronal loss or brain inflammation in the CSF of elderly subjects: 1) levels of the dopaminotrophic factors BDNF, GDNF, persephin, and neurturin, 2) levels of the proinflammatory factors TGFß1 and TGFß2; 3) activity of main antioxidant enzymes (catalases, glutathione-peroxidase, glutathione-reductase, glutathione-S-transferases, peroxirredoxins, and superoxide-dismutases), 4) ferritin content, antioxidant protein which reduces reactive free iron, and 5) antioxidant potential of the cerebrospinal fluid. ELISA and PAO tests were used. Subjects were also evaluated clinically, and the group of old subjects with mild cognitive impairment was studied separately. The findings indicate that normal elderly CSF is devoid of changes in either dopaminotrophic or proinflammatory factors. The antioxidant efficacy is slightly reduced with normal aging, through a reduction of glutathione-S-transferase activity in people older than 74 years (p < 0.05). However old people with mild cognitive impairment show reduced BDNF levels, and stronger signs of oxidative stress such as low antioxidant potential and glutathione-S-transferase activity (p < 0.05). To sum up, the present study demonstrates that, in CSF of normal senescence, dopaminotrophic factors and proinflammatory TGF-family ligands are not affected, and antioxidant efficacy is slightly reduced. CSF of elderly subjects with mild cognitive impairment shows more oxidative and trophic changes that are characterized by reduction of BDNF content, glutathione-S-transferase activity, and antioxidant potential.

The transient receptor potential vanilloid-1 is localized at excitatory synapses in the mouse dentate gyrus.

The transient receptor potential vanilloid type 1 (TRPV1) is a non-selective cation channel that plays an important role in pain perception and modulates neurotransmitter release and synaptic plasticity in the brain. TRPV1 function must lay on its anatomical distribution in the peripheral and central nervous system regions involved in the physiological roles of the channel. However, the anatomical localization of TRPV1 is well established in the periphery, but in the brain it is a matter of debate. While some studies support the presence of TRPV1 in several brain regions, recent evidences suggest a restricted distribution of the channel in the central nervous system. To investigate to what extent central TRPV1 function stands on a precise brain distribution of the channel, we examined the mouse hippocampal dentate molecular layer (ML) where TRPV1 mediates long-term synaptic plasticity. Using pre-embedding immunocytochemistry for high resolution electron microscopy, we show that TRPV1 immunoparticles are highly concentrated in postsynaptic dendritic spines to asymmetric perforant path synapses in the outer 2/3 of the ML. However, TRPV1 is poorly expressed at the excitatory hilar mossy cell synapses in the inner 1/3 of this layer. Importantly, the TRPV1 pattern distribution disappeared in the ML of TRPV1-knockout mice. Taken together, these findings support the notion of the presence of TRPV1 in a brain region where the channel has been shown to have a functional role, such as the perforant path synapses in the hippocampal dentate ML.

Effects of acute and repeated cocaine on markers for neural plasticity within the mesolimbic system in rats.

RATIONALE: Repeated cocaine is known to induce morphological changes in dopaminergic circuits that are known to participate on cocaine-induced addictive changes. OBJECTIVE: The objective of the present study was to discern if acute or repeated regimens of daily cocaine (10 mg/kg) lead to reliable changes in the expression of some protein markers for neural plasticity such as synaptophysin, p21-Arc, alpha-tubulin (α-tubulin), and stathmin, in the mesolimbic dopaminergic circuit. Well-known changes in tyrosine hydroxylase and protein kinase A were used for confirming biochemical effects of repeated cocaine. Animals were subjected to three treatments: acute injection, 3-day injections, or sensitizing cocaine during 3 days followed by challenging doses at days 8 and 18. RESULTS: The findings revealed that sensitizing regimen of cocaine increases stathmin levels within the nucleus accumbens at day 18 of treatment, not day 8, without changes of synaptophysin, p21-Arc, or α-tubulin. This neural plasticity change seems not to be related to the development of motor sensitization. Other neural regions such as prefrontal cortex, dorsal striatum, and ventral tegmental area were not found to be affected. Repeated cocaine led to well-known short-term augmentation of tyrosine-hydroxylase and protein kinase A expressions in the nucleus accumbens, as well as maintained upregulation of tyrosine hydroxylase in the ventral tegmental area. CONCLUSIONS: As stathmin is an important regulatory protein of microtubule dynamics, this protein change would be linked to morphological changes after repeated cocaine. It was confirmed that upregulation of tyrosine hydroxylase within the ventral tegmental area may participate on the development of motor sensitization.

The systemic administration of oleoylethanolamide exerts neuroprotection of the nigrostriatal system in experimental Parkinsonism.

Oleoylethanolamide (OEA) is an agonist of the peroxisome proliferator-activated receptor α (PPARα) and has been described to exhibit neuroprotective properties when administered locally in animal models of several neurological disorder models, including stroke and Parkinson's disease. However, there is little information regarding the effectiveness of systemic administration of OEA on Parkinson's disease. In the present study, OEA-mediated neuroprotection has been tested on in vivo and in vitro models of 6-hydroxydopamine (6-OH-DA)-induced degeneration. The in vivo model was based on the intrastriatal infusion of the neurotoxin 6-OH-DA, which generates Parkinsonian symptoms. Rats were treated 2 h before and after the 6-OH-DA treatment with systemic OEA (0.5, 1, and 5 mg/kg). The Parkinsonian symptoms were evaluated at 1 and 4 wk after the development of lesions. The functional status of the nigrostriatal system was studied through tyrosine-hydroxylase (TH) and hemeoxygenase-1 (HO-1, oxidation marker) immunostaining as well as by monitoring the synaptophysin content. In vitro cell cultures were also treated with OEA and 6-OH-DA. As expected, our results revealed 6-OH-DA induced neurotoxicity and behavioural deficits; however, these alterations were less severe in the animals treated with the highest dose of OEA (5 mg/kg). 6-OH-DA administration significantly reduced the striatal TH-immunoreactivity (ir) density, synaptophysin expression, and the number of nigral TH-ir neurons. Moreover, 6-OH-DA enhanced striatal HO-1 content, which was blocked by OEA (5 mg/kg). In vitro, 0.5 and 1 µM of OEA exerted significant neuroprotection on cultured nigral neurons. These effects were abolished after blocking PPARα with the selective antagonist GW6471. In conclusion, systemic OEA protects the nigrostriatal circuit from 6-OH-DA-induced neurotoxicity through a PPARα-dependent mechanism.

Oleoylethanolamide dose-dependently attenuates cocaine-induced behaviours through a PPARα receptor-independent mechanism.

Oleoylethanolamide (OEA) is an acylethanolamide that acts as an agonist of nuclear peroxisome proliferator-activated receptor alpha (PPARα) to exert their biological functions, which include the regulation of appetite and metabolism. Increasing evidence also suggests that OEA may participate in the control of reward-related behaviours. However, direct experimental evidence for the role of the OEA-PPARα receptor interaction in drug-mediated behaviours, such as cocaine-induced behavioural phenotypes, is lacking. The present study explored the role of OEA and its receptor PPARα on the psychomotor and rewarding responsiveness to cocaine using behavioural tests indicative of core components of addiction. We found that acute administration of OEA (1, 5 or 20 mg/kg, i.p.) reduced spontaneous locomotor activity and attenuated psychomotor activation induced by cocaine (20 mg/kg) in C57Bl/6 mice. However, PPARα receptor knockout mice showed normal sensitization, although OEA was capable of reducing behavioural sensitization with fewer efficacies. Furthermore, conditioned place preference and reinstatement to cocaine were intact in these mice. Our results indicate that PPARα receptor does not play a critical, if any, role in mediating short- and long-term psychomotor and rewarding responsiveness to cocaine. However, further research is needed for the identification of the targets of OEA for its inhibitory action on cocaine-mediated responses.

Alteration of neuropathic and visceral pain in female C57BL/6J mice lacking the PPAR-α gene.

RATIONALE: Peroxisome proliferator-activated receptors (PPARs) participate in the control of chronic neuropathic and inflammatory pain, and these receptors could play a role on acute pain. OBJECTIVES: We used null (PPAR-α -/-) and wild-type female mice and the PPAR-α blocker GW6471 to evaluate (1) the role of PPAR-α on neuropathic pain, (2) the involvement of PPAR-α on visceral and acute thermal nociception, and (3) tissue levels of pro-inflammatory factors. METHODS: Neuropathic pain was induced by sciatic nerve ligature. Acute thermal nociception was evaluated through hot-plate, tail-immersion, and writhing tests. The pro-inflammatory factors nitric oxide, TNF-α, and interleukins-1ß and -3 were measured. RESULTS: Regarding neuropathic pain, higher sensitivity to thermal and mechanical non-noxious and noxious stimuli was observed in mice lacking PPAR-α. Cold and mechanical allodynia and heat hyperalgesia were augmented in null mice. With respect to visceral nociception, writhes after acetic acid were enhanced in mutant mice. Although basal thermal sensitivity was enhanced in PPAR-α -/- mice, cutaneous thermal nociception did not differ between genotypes. Blockade of PPAR-α was devoid of effects on acute thermal and writhing tests. Finally, nerve ligature enhanced pro-inflammatory factors in plantar tissue, levels being higher in null mice. No changes in pro-inflammatory factors were observed in the hot-plate test. CONCLUSIONS: Genetic ablation of PPAR-α is involved in neuropathic and visceral nociception. Lack of PPAR-α is not involved in acute thermal pain, but it is involved in basal thermal reaction. Changes are biological adaptations to receptor deletion because blockade of PPAR-α does not affect inflammatory pain or thermal reactions.

Sensitization to cocaine is inhibited after intra-accumbal GR103691 or rimonabant, but it is enhanced after co-infusion indicating functional interaction between accumbens D(3) and CB1 receptors.

RATIONALE: Dopamine D(3) receptors and cannabinoid CB(1) receptors are both expressed in the nucleus accumbens, and they have been involved in motor sensitization to cocaine. The objectives were: (1) to study the effects of blockade of these receptors on sensitization to repeated cocaine, by using GR103691, D(3) receptor blocker, and rimonabant, CB(1) receptor ligand, and (2) to discern if both receptors interact by co-infusing them. MATERIALS AND METHODS: Cocaine (10 mg/kg) was injected daily for 3 days (induction phase) and later on day 8 (expression phase), and locomotor activity was measured during 2 h after cocaine. GR103691 and rimonabant were bilaterally injected (0.5 µl volume of each infusion) in the nucleus accumbens through cannulae (GR103691, 0, 4.85, and 9.7 µg/µl; rimonabant, 0, 0.5, and 1.5 µg/µl), before cocaine, during either induction or expression phases of sensitization. RESULTS: The findings indicated that sensitizing effects of cocaine were abolished after D(3) receptor blocking during both induction and expression phases, as well as rimonabant infusion during the expression (not induction) phase. A functional interaction between both receptors was also observed, because if GR103691 was injected during induction and rimonabant during expression, sensitizing effects of cocaine were observed to be normal or further enhanced. CONCLUSION: Dopamine D(3) receptors within the nucleus accumbens are critical for the development and consolidation of sensitization, and cannabinoid CB(1) receptors are critical for the expression of sensitization. Co-blockade of D(3) and CB(1) receptors exert opposite effects to blockade of these receptors separately, revealing the existence of a functional interaction between them.

Antiparkinsonian trophic action of glial cell line-derived neurotrophic factor and transforming growth factor ß1 is enhanced after co-infusion in rats.

The objective was to analyze functional effects of the combination of GDNF and TGF-ß1 in the retrograde model of Parkinsonism in rats, based on the intrastriatal infusion of 6-hydroxydopamine, which leads to protracted and progressive cell death in the substantia nigra. Hemiparkinsonian rats were implanted with osmotic minipumps 2 months after striatal lesion, pumps delivering GDNF alone (10 ng/day), TGF-ß1 alone (2 ng/day), or a GDNF and TGF-ß1 combination. The findings confirmed that GDNF alone has potent dopaminotrophic effects but they also revealed, for the first time, that GDNF and TGF-ß1 co-infusion led to stronger trophic effects relative to the infusion of GDNF alone. TGF-ß1 allowed further reducing dopamine receptor hypersensitivity, and potentiated GDNF-mediated effects. This cooperation could be accounted for by the recruitment of GFRα1 on striatal membranes, and by enhanced expression and activation of TH through augmented pSer31TH and pSer40TH. Co-infusion induced striatal sprouting, as revealed by augmentation of p21-Arc, stathmin, and synaptophysin, and led to a reliable recovery of phenotypic expression of TH in surviving nigral neurons. Functional recovery and improvement of TH signal in the nigrostriatal system were long-lasting and sustained, remaining after cessation of trophic infusion.

Response to methadone maintenance treatment is associated with the MYOCD and GRM6 genes.

BACKGROUND: There is increasing interest in the pharmacogenetic basis for explaining differences between patients in treatment outcome among methadone-treated subjects. Most studies have focused on genetic polymorphisms related to methadone pharmacokinetics and, to a lesser extent, those genes implicated in the pharmacodynamics of methadone. OBJECTIVE: This study aimed to investigate the associations between response to methadone maintenance treatment (MMT) and polymorphisms in genes coding for the OPRM1 opioid receptor, the metabotropic glutamate receptors GRM6 and GRM8, the nuclear receptor NR4A2, the photolyase enzyme cryptochrome 1 (CRY1), and the transcription factor myocardin (MYOCD), which have previously been associated with the risk of opioid dependence disorder. METHODS: The study used an association, case-control design, conducted in the setting of an MMT program in a drug abuse outpatient center in Barcelona, Spain. We recruited 169 opioid-dependent patients (diagnosed according to the Diagnostic and Statistical Manual of Mental Disorders [4th Edition] criteria) receiving MMT. The inclusion criteria included Caucasian ethnicity, being enrolled in MMT for at least 6 months, and receiving a stable methadone dose for the previous 2 months. The exclusion criteria included language-related barriers, severe cognitive impairment, or any medical disorder that would interfere with the research assessments. Single nucleotide polymorphism (SNP) variants in several candidate genes and regions were genotyped: MYOCD (rs1714984), GRM8 (rs1034576), CRY1 (rs1861591), GRM6 (rs953741), OPRM1 (rs1074287), NR4A2 (rs1405735), and the intergenic variants rs965972 (1q31.2) and rs1867898 (2q21.2). MMT response status was assessed by the number of opioid-positive controls detected by random urinalysis in the previous 2 months. We used the chi-squared test and p-value for the allele frequencies of the eight SNPs in responders versus nonresponders, and multivariate logistic regression analyses to examine associations between genotypes in the responder and ronresponder groups under codominant, dominant, and recessive models of inheritance. RESULTS: A final sample of 116 opioid-dependent patients were included and classified as methadone responders (n = 83) and nonresponders (n = 33), according to illicit opioid use detection in random urinalysis. The responders and nonresponders showed similar demographic and clinical characteristics. All SNPs were in Hardy-Weinberg equilibrium. Subjects carrying the AA genotype at rs1861591 (CRY1; Chr 12: 105941056 G>A) had a higher risk of being nonresponders (odds ratio [OR] = 2.99; 95% CI 1.14, 7.85; p = 0.035), although this difference disappeared with multiple testing corrections. Patients carrying the A allele at rs1714984 (MYOCD; Chr 17: 12558425 G>A) had an increased risk of being nonresponders only if they were also carriers of the AG genotype at rs953741 (GRM6; Chr5: 178262451 A>G) [OR = 10.83; 95% CI 2.52, 46.66; p = 0.006]. CONCLUSIONS: A positive association was observed between response to methadone and two variants in the genes MYOCD and GRM6. A pharmacogenetic epistatic effect between SNPs in MYOCD and GRM6 appears to modulate inter-individual variations in MMT response.

Effects of the endogenous PPAR-alpha agonist, oleoylethanolamide on MDMA-induced cognitive deficits in mice.

MDMA (3,4-Methylenedioxymethamphetamine) is an amphetamine derivative widely used for recreational purposes. We have recently shown that repeated treatment with high doses of MDMA-induced impairments in the acquisition and recall of an active avoidance task in mice. In this study, we examined whether the endogenous peroxisome proliferator-activated receptor-alpha (PPAR-alpha) agonist, oleoylethanolamide (OEA) protects against these MDMA-induced deficits. Mice were pretreated twice a day with OEA (0, 5, and 25 mg/kg) 30 min before an injection of MDMA (30 mg/kg) or saline during four consecutive days. Twenty-four hours after the last treatment, animals were trained in an active avoidance task for two consecutive weeks. After a 5-day resting period, a recall session was performed. Mice treated with MDMA showed reduced learning and recall of the task when compared with saline-treated controls. OEA at 5 mg/kg ameliorated and at 25 mg/kg worsened this deficit. Dopamine transporter (DAT)-binding sites significantly decreased 4 days after the last MDMA administration and pretreatment with both doses of OEA prevented this effect. In immunohistochemical studies, coexpression of tyrosine-hydroxylase and PPAR-alpha receptors was observed in the striatum and substantia nigra pars compacta of mice. These results suggest that OEA administration can modulate the cognitive deficits induced by MDMA in a DAT-independent manner.