Jorge Diaz

Role of the dopamine D3 receptor in reactivity to cocaine‐associated cues in mice

Environmental stimuli previously associated with drug effects can acquire secondary reinforcing properties, able to maintain drug‐seeking behaviour or induce relapse. We have used a classical Pavlovian conditioning procedure to assess the role of the dopamine D3 receptor (D3R) in the expression of drug‐conditioned responses. Mice repeatedly receiving cocaine in a particular environment distinct from home‐cages displayed hyperlocomotion after subsequent exposure to the drug‐paired environment. Cocaine‐conditioned hyperactivity was inhibited by BPu2003897 or SB‐277011‐A, D3R‐selective partial agonist and antagonist, respectively. D3R gene‐targeted mice showed a trend towards an increase in cocaine cue‐conditioned hyperactivity. BPu2003897 had no effect on reactivity to neutral or aversive cues. Cocaine‐conditioned mice had increased levels of D3R mRNA and binding in the nucleus accumbens (NAc), and transcripts of brain‐derived neurotrophic factor (BDNF), a factor controlling D3R expression, in the ventral tegmental area (VTA). Cocaine had no effects on D3R or BDNF genes when administered in home‐cages. Cocaine cue‐conditioned c‐fos expression was found in cortical areas, notably in the somatosensory cortex, where it was inhibited by BPu2003897, and in several regions belonging or linked to the limbic system. In conditioned mice, BPu2003897 inhibited c‐fos expression in VTA and activated it in amygdala. These results demonstrate a modulation of reactivity to cocaine cues by the D3R, the expression of which is elevated in the NAc by the repeated association of drug effects with a particular context, through a BDNF‐dependent mechanism. D3R‐selective partial agonist or antagonist inhibit cocaine cue‐conditioned activity possibly by normalizing exacerbated D3R function in the NAc, but our results also point to a possible participation of a pathway involving the VTA, amygdala and somatosensory cortex.

Coexpression of dopamine D1 and D3 receptors in islands of Calleja and shell of nucleus accumbens of the rat: opposite and synergistic functional interactions

Using double in situ hybridization, we found extensive coexpression of dopamine D1 and D3 receptor (D1R and D3R) mRNAs in neurons of the island of Calleja major (ICjM) and ventromedial shell of nucleus accumbens (ShV), respectively. Thus, at least 79 and 63% of D3R mRNA‐expressing neurons in ICjM and ShV also expressed the D1R mRNA. Coexpression of D1R and D3R mRNAs was found to occur in substance P (SP) mRNA‐expressing neurons in both areas, suggesting SP mRNA as a marker of the activity of coexpressing neurons. Administration of SKF 38393, a D1R receptor agonist, increased c‐fos mRNA in ICjM, whereas administration of quinpirole, a D2R/D3R agonist, decreased it; SCH 23390, a D1R antagonist and nafadotride, a preferential D3R antagonist, given alone, had effects opposite to those of the corresponding agonists. These data indicate that basal c‐fos expression in ICjM is maintained by endogenous dopamine acting tonically upon two receptor subtypes subserving opposite effects on the same cell. However, in ShV, whereas SKF 38393 also increased c‐fos mRNA, quinpirole had no effect, a difference presumably reflecting the lower fraction of neurons coexpressing D1R and D3R in this area. In contrast, in ShV from reserpine‐treated rats, SKF 38393 increased SP mRNA and quinpirole potentiated this effect. These contrasting interactions of D1R‐ and D3R‐mediated signalling events, i.e. in either opposite or synergistic directions, most likely occurring at the single cell level, may serve to increase the dopamine response threshold of the target cells in ICjM and to maintain a strong tonic activity of ShV neurons.

Opposing roles for dopamine D2 and D3 receptors on neurotensin mRNA expression in nucleus accumbens.

Using in situ hybridization histochemistry in rat nucleus accumbens, we show that the dopamine D3 receptor mRNA is expressed in the ventromedial part of the shell subdivision, where its gross distribution matches that of neurotensin mRNA. In addition, hybridization studies at the cellullar level show that a large fraction of the neurotensin neurons co‐express the D3 receptor mRNA in this restricted area. In contrast, the dopamine D2 receptor mRNA is expressed mainly in the core and marginally in the shell, at the level of the cone. In rats treated by haloperidol and sulpiride, two D2‐like receptor antagonists, but not by SCH 23390, a D1‐like receptor antagonist, proneurotensin mRNA was increased in the D2 receptor mRNA‐rich areas but decreased in the D3 receptor mRNA‐rich areas. This suggests that the D2 and D3 receptors control neurotensin mRNA expression negatively and positively, respectively.

Direct and indirect interactions of the dopamine D3 receptor with glutamate pathways: implications for the treatment of schizophrenia

This article, based on original data as well as on previously reported preclinical and clinical data that are reviewed, describes direct and indirect interactions of the D3 receptor with N-methyl-d-aspartate receptor (NMDA) signaling and their functional consequences and therapeutic implications for schizophrenia. D3 receptor immunoreactivity at ultrastructural level with electron microscopy was identified at presumably glutamatergic, asymmetric synapses of the medium-sized spiny neurons of the nucleus accumbens. This finding supports the existence of a direct interaction of the D3 receptor with glutamate, in line with previously described interactions with NMDA signaling involving Ca2+/calmodulin-dependent protein kinase II at post-synaptic densities (Liu et al. 2009). Indirect interactions of the D3 receptor with glutamate could involve a negative control exerted by the D3 receptor on mesocortical dopamine neurons and the complex regulation of the glutamatergic pyramidal cells by dopamine in the prefrontal cortex. This could be exemplified here by the regulation of pyramidal cell activity in conditions of chronic NMDA receptor blockade with dizocilpine (MK-801). BP897, a D3 receptor-selective partial agonist, reversed the dysregulation of cortical c-fos mRNA expression and pyramidal cell hyperexcitability, as measured by paired-pulse electrophysiology. At the behavioral level, blockade of the D3 receptor, by known D3 receptor antagonists or the novel D3 receptor-selective antagonist F17141, produces antipsychotic-like effects in reversing hyperactivity and social interaction deficits induced by NMDA receptor blockade by MK-801 in mice. The glutamate–D3 receptor interactions described here offer a conceptual framework for developing new D3 receptor-selective drugs, which may appear as an original, efficacious, and safe way to potentially indirectly target glutamate in schizophrenia.

Mice Lacking GPR88 Show Motor Deficit, Improved Spatial Learning, and Low Anxiety Reversed by Delta Opioid Antagonist

BACKGROUNDnGPR88 is an orphan G protein coupled receptor highly enriched in the striatum, and previous studies have focused on GPR88 function in striatal physiology. The receptor is also expressed in other brain areas, and here we examined whether GPR88 function extends beyond striatal-mediated responses.nnnMETHODSnWe created Gpr88 knockout mice and examined both striatal and extrastriatal regions at molecular and cellular levels. We also tested striatum-, hippocampus-, and amygdala-dependent behaviors in Gpr88(-/-) mice using extensive behavioral testing.nnnRESULTSnWe found increased G protein coupling for delta opioid receptor (DOR) and mu opioid, but not other Gi/o coupled receptors, in the striatum of Gpr88 knockout mice. We also found modifications in gene transcription, dopamine and serotonin contents, and dendritic morphology inside and outside the striatum. Behavioral testing confirmed striatal deficits (hyperactivity, stereotypies, motor impairment in rotarod). In addition, mutant mice performed better in spatial tasks dependent on hippocampus (Y-maze, novel object recognition, dual solution cross-maze) and also showed markedly reduced levels of anxiety (elevated plus maze, marble burying, novelty suppressed feeding). Strikingly, chronic blockade of DOR using naltrindole partially improved motor coordination and normalized spatial navigation and anxiety of Gpr88(-/-) mice.nnnCONCLUSIONSnWe demonstrate that GPR88 is implicated in a large repertoire of behavioral responses that engage motor activity, spatial learning, and emotional processing. Our data also reveal functional antagonism between GPR88 and DOR activities in vivo. The therapeutic potential of GPR88 therefore extends to cognitive and anxiety disorders, possibly in interaction with other receptor systems.

Subapical Localization of the Dopamine D3 Receptor in Proximal Tubules of the Rat Kidney

The dopamine D3 receptor (D3R), intensively studied in neuroscience, also plays an important role in the regulation of renal and cardiovascular function. In contrast to functional findings, less information is available on its localization in the kidney. Neither RT-PCR studies nor radioligand binding assays are suitable to selectively determine the distribution of renal D3R at the level of cellular or even subcellular structures. We studied the renal D3R distribution in Sprague-Dawley rats by a polyclonal antiserum directed against an epitope in the third intracytoplasmic loop. D3R immunoreactivity was detected by indirect immunofluorescence and confocal laser scanning microscopy. D3R staining was confined to the renal cortex and occurred in proximal convoluted tubules near or in direct connection with the urinary pole of the glomeruli. The fluorescent spots were restricted to the subapical portion of the proximal tubular cells. Double staining with the F-actin marker phalloidin revealed a localization of the D3R below the brush border region. However, staining by anti-P1/p2-adaptins, recognizing clathrin-coated compartments, did not correspond to the distribution of the D3R signal. This is the first description of a D3R accumulation in a cytoplasmic pool in the kidney, probably corresponding to a recycling mechanism or storage compartment.

Aging, but not tau pathology, impacts olfactory performances and somatostatin systems in THY-Tau22 mice

Somatostatin (SOM) cortical levels decline in Alzheimers disease (AD) in correlation with cognitive impairment severity, the latter being closely related to the presence of neurofibrillary tangles. Impaired olfaction is another hallmark of AD tightly related to tau pathology in the olfactory pathways. Recent studies showed that SOM modulates olfactory processing, suggesting that alterations in SOM levels participate to olfactory deficits in AD. Herein, we first observed that human olfactory peduncle and cortex are enriched in SOM cells and fibers, in aged postmortem brains. Then, the possible link between SOM alterations and olfactory deficits was evaluated by exploring the impact of age and tau hyperphosphorylation on olfactory SOM networks and behavioral performances in THY-Tau22 mice, a tauopathy transgenic model. Distinct molecular repertoires of SOM peptide and receptors were associated to sensory or cortical olfactory processing structures. Aging mainly affected SOM neurotransmission in piriform and entorhinal cortex in wild-type mice, although olfactory performances decreased. However, no further olfactory impairment was evidenced in THY-Tau22 mice until 12 months although tau pathology early affected olfactory cortical structures. Thus, tau hyperphosphorylation per se has a limited impact on olfactory performances in THY-Tau22 mice.

Yif1B Is Involved in the Anterograde Traffic Pathway and the Golgi Architecture

Yif1B is an intracellular membrane‐bound protein belonging to the Yip family, shown previously to control serotonin 5‐HT1A receptor targeting to dendrites. Because some Yip proteins are involved in the intracellular traffic between the ER and the Golgi, here we investigated the precise localization of Yif1B in HeLa cells. We found that Yif1B is not resident into the Golgi, but rather belongs to the IC compartment. After analyzing the role of Yif1B in protein transport, we showed that the traffic of the VSVG protein marker was accelerated in Yif1B depleted HeLa cells, as well as in hippocampal neurons from Yif1B KO mice. Conversely, Yif1B depletion in HeLa cells did not change the retrograde traffic of ShTx. Interestingly, in long term depletion of Yif1B as in Yif1B KO mice, we observed a disorganized Golgi architecture in CA1 pyramidal hippocampal neurons, which was confirmed by electron microscopy. However, because short term depletion of Yif1B did not change Golgi structure, it is likely that the implication of Yif1B in anterograde traffic does not rely on its role in structural organization of the Golgi apparatus, but rather on its shuttling between the ER, the IC and the Golgi compartments.

BIN1 genetic risk factor for Alzheimer is sufficient to induce early structural tract alterations in entorhinal cortex-dentate gyrus pathway and related hippocampal multi-scale impairments

Late-onset Alzheimer Disease (LOAD) is the most common form of dementia and one of the most challenging diseases of modern society 1. Understanding the preclinical stages of AD that begins in the brain at least 2-3 decades before evidence of episodic memory defects in patients is pivotal for the design of successful approaches to delay or reverse the transition from normal brain function to cognitive impairments. Our working hypothesis is that LOAD genetic risk factors can be sufficient to generate early phenotypical changes before any changes in either Abeta or Tau. To test this hypothesis, we generated an hBIN1 mouse model based on the human BIN1 gene overexpression that we found in post-mortem brain samples from LOAD patients. BIN1 is the second important risk factor for AD, following the APOE gene 2. We identified co-deregulated gene repertoires common to both 7-week mouse hippocampus sub-regions and post-mortem brain samples from LOAD patients, demonstrating the validity of this hBIN1 model. We evidenced an early phenotype of neurodegeneration starting at 3 months with structural impairment fiber pathways quantified by high resolution (17.2T) (MRI-DTI) and related functional impacts. We found structural changes in entorhinal cortex-dentate gyrus (EC-DG) pathway known to be the earliest brain region impacted in LOAD 3–6. Similarly, the function of this pathway was impaired both in vitro and in vivo, with the changes in spine density and dendritic simplification of DG neurons, impaired EC-DG long-term potentiation (LTP) and behavioral deficits linked to object recognition episodic memory. As expected for a neurodegenerative model, we evidenced the progression of dysfunction at the morphological, functional and behavioral levels with age. Structural spreading involved impairment of fibers in somatosensory and temporal associative cortexes at month 15. Functional and behavioral spreading was characterized by impact on pattern separation of spatial episodic memory. Moreover, this neurodegeneration occurred without any detectable changes in Aβ42 and tau. Overall, these data show the possibility to identify a repertoire of molecular changes occurring both in patients and in hBIN1 mice and whose further manipulation can be instrumental to rescue or delay episodic memory defects.