Distribution of tritium labeled 12(S) hydroxy-eicosatetraenoic acid (12-HETE) in the rat.

The in vivo metabolism of 12-(S)-Hydroxy-eicosatetraenoic acid (12-HETE), the end-lipoxygenase product of arachidonic acid in platelets, has been investigated in the rat. Fifty microcuries of 5,6-[3H]-12-HETE (50 Ci/mmol) were injected to anesthetized rats and the radioactivity was followed in plasma. At the end of the experiment, various organs of the animal were removed and the radioactivity attached to them was determined. The label of the plasma plateaued to approximately one third of the initial radioactivity ten minutes after the injection. Among the various organs tested (brain, heart, intestine, kidney, liver, lungs, spleen, testis/uterus) the kidney was far the most active to accumulate 12-HETE and/or its labeled metabolites, and no radioactivity could be detected in urine during the course of the experiment. The analysis of lipid extracts from the various tissues revealed that 12-HETE was not accumulating in its unesterified form but was likely bound to phospholipids. We conclude that, although the label providing from the initial 12-HETE did not completely disappear from plasma, circulating 12-HETE cannot be considered as a circulating marker of cell activation.     

Elongation and desaturation of arachidonic and eicosapentaenoic acids in rat liver. Effect of clofibrate feeding.

The fatty acid elongation-desaturation ability of 5,8,11,14-eicosatetraenoic (20:4(n-6)) and 5,8,11,14,17-eicosapentaenoic (20:5(n-3)) acids was determined in both liver microsomal and light mitochondrial (rich in peroxisomes) fractions of untreated and clofibrate treated rats. The elongation and the subsequent desaturation steps were performed in the corresponding favorable media. 20:5(n-3) elongation was about 2-times more extensive than that of 20:4(n-6). Clofibrate feeding for 10 days resulted in a marked decrease in the elongation rate with the two substrates, while the delta 4 desaturation rate was increased. There were small differences in the elongation rate between the microsomal and light mitochondrial fractions, however, the relative delta 4 desaturation rate was higher in the light mitochondrial fraction than microsomes.     

Behavioural Profiles in Captive-Bred Cynomolgus Macaques: Towards Monkey Models of Mental Disorders?

Background

To date, experimental and preclinical studies on neuropsychiatric conditions have almost exclusively been performed in experimentally-induced animal models and have only rarely relied upon an ethological approach where animals have been observed in more naturalistic settings. The laboratory species of choice has been the rodent while the potential of more closely-related non-human primates have remained largely underexplored.

Methods

The present study, therefore, aimed at investigating the possible existence of spontaneous atypical/abnormal behaviours displayed by 40 cynomolgus macaques in captive conditions using an unbiased ethological scan-sampling analysis followed by multifactorial correspondence analysis and a hierarchical clustering.

Results

The study identified five distinct profiles (groups A to E) that significantly differed on several behaviours, body postures, body orientations, gaze directions and locations in the cage environment. We suggest that animals from the low n groups (D and E) present depressive-like and anxious-like symptoms, reminiscent of depressive and generalized anxiety disorders. Inter-individual differences were highlighted through unbiased ethological observations of spontaneous behaviours and associated parameters, although these were not associated with differences in plasma or cerebrospinal fluid levels of either stress-related hormones or monoamines, i.e. in accordance with the human situation.

Conclusions

No interventional behavioural testing was required to discriminate between 3 typical and 2 atypical ethologically-defined behavioural profiles, reminiscent of certain depressive-like and anxiety-like symptoms. The use of unbiased behavioural observations might, thus, allow the identification of animal models of human mental/behavioural disorders and their most appropriate control groups.     

Lysosomal dysfunction in Parkinson disease: ATP13A2 gets into the groove

Mutations in ATP13A2 (PARK9) cause an autosomal recessive form of early-onset parkinsonism with pyramidal degeneration and dementia called Kufor-Rakeb Syndrome (KRS). The ATP13A2 gene encodes a transmembrane lysosomal P5-type ATPase (ATP13A2) whose physiological function in mammalian cells, and hence its potential role in Parkinson disease (PD), remains elusive. In this context, we have recently shown that KRS-linked mutations in ATP13A2 leads to several lysosomal alterations in ATP13A2 KRS patient-derived fibroblasts, including impaired lysosomal acidification, decreased proteolytic processing of lysosomal enzymes, reduced degradation of lysosomal substrates and diminished lysosomal-mediated clearance of autophagosomes (AP). Similar alterations are observed in stable ATP13A2-knockdown dopaminergic cell lines, which are associated with cell death. Restoration of ATP13A2 levels in ATP13A2-mutant/depleted cells is able to restore lysosomal function and attenuate cell death. Relevant to PD, we have determined that ATP13A2 levels are decreased in dopaminergic nigral neurons from sporadic PD patients. Interestingly in these patients, the main signal of ATP13A2 is detected in the Lewy bodies. Our results unravel an instrumental role of ATP13A2 in lysosomal function and in cell viability. Altogether, our results validate ATP13A2 as a likely therapeutic target against PD degeneration.     

Neuroanatomical study of the A11 diencephalospinal pathway in the non-human primate

Background

The A11 diencephalospinal pathway is crucial for sensorimotor integration and pain control at the spinal cord level. When disrupted, it is thought to be involved in numerous painful conditions such as restless legs syndrome and migraine. Its anatomical organization, however, remains largely unknown in the non-human primate (NHP). We therefore characterized the anatomy of this pathway in the NHP.

Methods and Findings

In situ hybridization of spinal dopamine receptors showed that D1 receptor mRNA is absent while D2 and D5 receptor mRNAs are mainly expressed in the dorsal horn and D3 receptor mRNA in both the dorsal and ventral horns. Unilateral injections of the retrograde tracer Fluoro-Gold (FG) into the cervical spinal enlargement labeled A11 hypothalamic neurons quasi-exclusively among dopamine areas. Detailed immunohistochemical analysis suggested that these FG-labeled A11 neurons are tyrosine hydroxylase-positive but dopa-decarboxylase and dopamine transporter-negative, suggestive of a L-DOPAergic nucleus. Stereological cell count of A11 neurons revealed that this group is composed by 4002±501 neurons per side. A 1-methyl-4-phenyl-1, 2, 3, 6-tetrahydropyridine (MPTP) intoxication with subsequent development of a parkinsonian syndrome produced a 50% neuronal cell loss in the A11 group.

Conclusion

The diencephalic A11 area could be the major source of L-DOPA in the NHP spinal cord, where it may play a role in the modulation of sensorimotor integration through D2 and D3 receptors either directly or indirectly via dopamine formation in spinal dopa-decarboxylase-positives cells.     

Acidic nanoparticles protect against α‐synuclein‐induced neurodegeneration through the restoration of lysosomal function

Parkinson''s disease (PD) is an age‐related neurodegenerative disorder characterized by the loss of dopaminergic neurons in the substantia nigra, associated with the accumulation of misfolded α‐synuclein and lysosomal impairment, two events deemed interconnected. Protein aggregation is linked to defects in degradation systems such as the autophagy‐lysosomal pathway, while lysosomal dysfunction is partly related to compromised acidification. We have recently proven that acidic nanoparticles (aNPs) can re‐acidify lysosomes and ameliorate neurotoxin‐mediated dopaminergic neurodegeneration in mice. However, no lysosome‐targeted approach has yet been tested in synucleinopathy models in vivo. Here, we show that aNPs increase α‐synuclein degradation through enhancing lysosomal activity in vitro. We further demonstrate in vivo that aNPs protect nigral dopaminergic neurons from cell death, ameliorate α‐synuclein pathology, and restore lysosomal function in mice injected with PD patient‐derived Lewy body extracts carrying toxic α‐synuclein aggregates. Our results support lysosomal re‐acidification as a disease‐modifying strategy for the treatment of PD and other age‐related proteinopathies.     

High frequency stimulation of the entopeduncular nucleus has no effect on striatal dopaminergic transmission

High frequency stimulation (HFS) of the subthalamic nucleus (STN) is thought to be superior to stimulation of the internal pallidum (GPi) in alleviating symptoms of Parkinson's disease (PD). However, preliminary controlled studies comparing the effectiveness of both targets have not found significant differences in the improvement of parkinsonian symptoms, but have shown that STN stimulation allows a dramatic decrease in dopaminergic medication. We have previously shown that STN-HFS increases striatal extracellular dopamine (DA) metabolites, but not DA, in both naive and 6-hydroxydopamine (6-OHDA)-lesioned rats, whereas stimulation of the entopeduncular nucleus (EP), the rodent equivalent of the internal pallidum, does not affect DA or metabolite levels. Intriguingly, STN-HFS increases striatal DA release after inhibition of DA reuptake or metabolism, suggesting that this observation may have been obscured in non-drug treated animals by rapid and effective DA reuptake. Since STN-HFS further enhances DA metabolism after DA reuptake inhibition or depletion it has been proposed that STN-HFS increases both, striatal DA release and metabolism, independently. Therefore, the present study assesses the impact of EP-HFS on striatal DA release and metabolism in normal rats after inhibition of DA reuptake or metabolism, using microdialysis. In summary, our data demonstrate that, contrary to STN stimulation, EP-HFS has no effect on striatal DA release and metabolism. Thus, the present study provides a partial explanation for the reported clinical differences, and experimental evidence for differential mechanisms of action between HFS of the internal pallidum and the STN, that are most likely related to differences in functional anatomy.     

CD8+ cytotoxic T lymphocyte activation by soluble major histocompatibility complex-peptide dimers

CD8+ cytotoxic T lymphocyte (CTL) can recognize and kill target cells that express only a few cognate major histocompatibility complex class I-peptide (pMHC) complexes. To better understand the molecular basis of this sensitive recognition process, we studied dimeric pMHC complexes containing linkers of different lengths. Although dimers containing short (10-30-A) linkers efficiently bound to and triggered intracellular calcium mobilization and phosphorylation in cloned CTL, dimers containing long linkers (> or = 80 A) did not. Based on this and on fluorescence resonance energy transfer experiments, we describe a dimeric binding mode in which two T cell receptors engage in an anti-parallel fashion two pMHC complexes facing each other with their constant domains. This binding mode allows integration of diverse low affinity interactions, which increases the overall binding and, hence, the sensitivity of antigen recognition. In proof of this, we demonstrated that pMHC dimers containing one agonist and one null ligand efficiently activate CTL, corroborating the importance of endogenous pMHC complexes in antigen recognition.