Aripiprazole protects cortical neurons from glutamate toxicity.

Neurodegeneration is thought to be a component of schizophrenia pathology, and some antipsychotics appear to slow degenerative changes in patients. Aripiprazole, the first partial dopamine D(2) receptor agonist approved for the treatment of schizophrenia, is suggested to be neuroprotective based on non-clinical studies using transformed cell lines and in vivo stress and lesion paradigms. However, aripiprazole-induced neuroprotection has not been studied in a neuronal glutamate toxicity assay, which may model aspects of neurodegeneration occurring in schizophrenia. This study examined whether therapeutically relevant concentrations of aripiprazole protect rat embryonic cortical neurons from glutamate toxicity in biochemical and high-content imaging assays. Aripiprazole inhibited glutamate-induced neurotoxicity by 40% in a 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl-tetrazolium bromide (MTT) assay, in contrast to risperidone and olanzapine, which had little neuroprotective activity. This neuroprotective effect of aripiprazole was not mediated by the activation of serotonin 5-HT(1A) or dopamine D(2) receptors, Akt or glycogen-synthase kinase-3ß signaling (GSK-3ß), or through the inhibition of poly-ADP ribose polymerase (PARP). Further experiments are required to determine the biochemical nature of aripiprazole-induced neuroprotection and whether any such activity might have clinical relevance.

In vitro functional characteristics of dopamine D2 receptor partial agonists in second and third messenger-based assays of cloned human dopamine D2Long receptor signalling.

Aripiprazole, (+)terguride, OPC-4392 and (-)3-PPP have been classified as dopamine D(2) receptor partial agonists based largely on their activity in second messenger-based assays of dopamine D(2) receptor signalling. Nevertheless, signal transduction amplification might result in these compounds behaving as dopamine D(2) receptor full agonists at a more downstream level of signalling. We compared the intrinsic activity (E(max), expressed as a percentage of the maximal effect of dopamine) of aripiprazole, (+)terguride, OPC-4392 and (-)3-PPP using second (calcium (Ca(2+)) mobilization) and third (extracellular signal-regulated kinase 2 (ERK2) phosphoprotein expression) messenger readouts of cloned human dopamine D(2long) (hD(2L)) receptor signalling in CHO cells. These compounds were all less potent and displayed lower intrinsic activity in the Ca(2+) assay (aripiprazole = 24.3%, (+)terguride = 56.9%, OPC-4392 = 58.6% and (-)3-PPP = 75.1%), and aripiprazole (E(max) = 54.5%) displayed a substantially lower intrinsic activity than (+)terguride (E(max) = 92.3%), OPC-4392 (E(max) = 93.1%) and (-)3-PPP (E(max) = 101.1%) in the more downstream-based ERK2 phosphoprotein expression assay. These drug effects on Ca(2+) mobilization and ERK2 phosphoprotein expression were mediated through dopamine hD(2L) receptors, as they all were blocked by (-)raclopride, whereas (-)raclopride and other dopamine D(2) receptor antagonists (haloperidol, risperidone, ziprasidone, olanzapine, clozapine and quetiapine) were inactive on their own in both assays. These data are consistent with clinical evidence that only dopamine D(2) receptor partial agonists with a sufficiently low enough intrinsic activity will prove effective against the positive symptoms of schizophrenia, and also highlight the importance of using downstream-based assays in the discovery of novel D(2) receptor partial agonist therapeutics.

Dopamine D2 receptor partial agonists display differential or contrasting characteristics in membrane and cell-based assays of dopamine D2 receptor signaling.

Clinical evidence suggests that dopamine D(2) receptor partial agonists must have a sufficiently low intrinsic activity to be effective as antipsychotics. Here, we used dopamine D(2) receptor signaling assays to compare the in vitro functional characteristics of the antipsychotic aripiprazole with other dopamine D(2) receptor partial agonists (7-{3-[4-(2,3-dimethylphenyl)-piperazinyl]propoxy}-2(1H)-quinolinone [OPC-4392], (-)-3-(3-hydroxy-phenyl)-N-n-propylpiperidine [(-)3-PPP] and (+)terguride) and dopamine D(2) receptor antagonists. Aripiprazole and OPC-4392 were inactive in a guanosine-5'-O-(3-[(35)S]thio)-triphosphate ([(35)S]GTPgammaS) binding assay using Chinese Hamster Ovary (CHO) cell membranes expressing cloned human dopamine D(2Long) (hD(2L)) receptors, whereas (-)3-PPP and (+)terguride displayed low intrinsic activity. Aripiprazole also had no effect on [(35)S]GTPgammaS binding to CHO-hD(2L) cells, while OPC-4392, (-)3-PPP and (+)terguride were partial agonists. In contrast, aripiprazole, OPC-4392, (-)3-PPP, and (+)terguride were inactive in a [(35)S]GTPgammaS binding assay using rat striatal membranes. However, at a more downstream level of CHO-hD(2L) cell signalling, these drugs all behaved as dopamine hD(2L) receptor partial agonists, with aripiprazole displaying an intrinsic activity 2 to 3-fold lower (inhibition of forskolin-induced adenosine 3',5'-cyclic monophosphate accumulation) and almost half as high (enhancement of adenosine triphosphate-stimulated [(3)H]arachidonic acid release) as OPC-4392, (-)3-PPP and (+)terguride. Dopamine activity was blocked in each case by (-)raclopride, which was inactive on its own in every assay, as were the antipsychotics haloperidol, olanzapine, ziprasidone and clozapine. Together, these data, whilst preclinical in nature, are consistent with clinical evidence suggesting the favorable antipsychotic profile of aripiprazole, compared with the other clinically ineffective partial agonists, is dependent on its low intrinsic activity at dopamine D(2) receptors. This study also highlights the limitations of using [(35)S]GTPgammaS binding assays to identify dopamine D(2) receptor partial agonists.

In vitro biochemical evidence that the psychotomimetics phencyclidine, ketamine and dizocilpine (MK-801) are inactive at cloned human and rat dopamine D2 receptors.

Dopamine potently increased calcium mobilization in Chinese hamster ovary cells expressing human dopamine D2Long receptors (CHO-D2L cells), and increased guanosine-5'-O-(3-[35S]thio)-triphosphate binding to CHO-D2L cell and rat striatal membranes. These effects of dopamine were blocked by the dopamine D2 receptor antagonist (-)raclopride. In contrast to the findings of a recent controversial study, phencyclidine, ketamine and dizocilpine (MK-801) lacked dopamine D2 receptor full agonist, partial agonist and antagonist activity in these assays, suggesting their psychotomimetic effects, and activity in rodent models of schizophrenia, are associated with N-methyl-d-aspartate receptor blockade rather than a direct interaction with dopamine D2 receptors.