Fluoxetine, not donepezil, reverses anhedonia, cognitive dysfunctions and hippocampal proteome changes during repeated social defeat exposure.

While anhedonia is considered a core symptom of major depressive disorder (MDD), less attention has been paid to cognitive dysfunctions. We evaluated the behavioural and molecular effects of a selective serotonin re-uptake inhibitor (SSRI, fluoxetine) and an acetylcholinesterase inhibitor (AChEI, donepezil) on emotional-cognitive endophenotypes of depression and the hippocampal proteome. A chronic social defeat (SD) procedure was followed up by "reminder" sessions of direct and indirect SD. Anhedonia-related behaviour was assessed longitudinally by intracranial self-stimulation (ICSS). Cognitive dysfunction was analysed by an object recognition test (ORT) and extinction of fear memory. Tandem mass spectrometry (MSE) and protein-protein-interaction (PPI) network modelling were used to characterise the underlying biological processes of SD and SSRI/AChEI treatment. Independent selected reaction monitoring (SRM) was conducted for molecular validation. Repeated SD resulted in a stable increase of anhedonia-like behaviour as measured by ICSS. Fluoxetine treatment reversed this phenotype, whereas donepezil showed no effect. Fluoxetine improved recognition memory and inhibitory learning in a stressor-related context, whereas donepezil only improved fear extinction. MSE and PPI network analysis highlighted functional SD stress-related hippocampal proteome changes including reduced glutamatergic neurotransmission and learning processes, which were reversed by fluoxetine, but not by donepezil. SRM validation of molecular key players involved in these pathways confirmed the hypothesis that fluoxetine acts via increased AMPA receptor signalling and Ca2+-mediated neuroplasticity in the amelioration of stress-impaired reward processing and memory consolidation. Our study highlights molecular mediators of SD stress reversed by SSRI treatment, identifying potential viable future targets to improve cognitive dysfunctions in MDD patients.

Investigating the effect of bilateral amygdala lesions on fear conditioning and social interaction in the male Mongolian gerbil.

Identification of the selective neurokinin NK(1) receptor antagonist, 2-(R)-(1-(R)-3,5-Bis(trifluromethyl)phenylethoxy)-3-(S)-(4-fluoro)phenyl-4-(3-oxo-1,2,4-triazol-5yl)methylmor-phine (MK-869), as a novel therapeutic approach for anxiety/depression has led to increased use of the Mongolian gerbil in behavioural studies since the gerbil NK(1) receptor pharmacology is similar to human, but not rat or mouse. Within this species, foot tapping and immobility elicited by aversive conditioning, as well as social interaction have been shown to be sensitive to clinically used anxiolytic and antidepressant agents and also NK(1) receptor antagonists. The high levels of NK(1) receptor binding in the amygdala as well as preclinical studies demonstrating increased release of substance P and corresponding internalisation of NK(1) receptors in the basolateral amygdala in response to stressful stimuli suggest that the BLA may represent a potential site of action for NK(1) receptor antagonists in anxiety and/or depression. Therefore, in the current study, we assessed the effect of bilateral BLA lesions in male Mongolian gerbils on footshock-induced foot tapping and immobility, social interaction, and NK(1)-agonist-induced foot tapping. Lesioned gerbils exhibited reduced immobility time during fear conditioning, a non-significant reduction in immobility time when re-exposed to the conditioned stimulus (CS) 24 h later, and increased social interaction in the gerbil social interaction task. In contrast, BLA lesions had no effect on NK(1)-agonist-induced foot tapping. These data provide further support that the gerbil BLA is a potential site for NK(1) receptor antagonists to attenuate anxiety-related behaviours.

Donepezil reverses a mnemonic deficit produced by scopolamine but not by perforant path lesion or transient cerebral ischaemia.

The purpose of these studies were threefold. Firstly, to further characterize the effect of perforant path transection on a test of short-term memory: delayed matching (or nonmatching)-to-position [D(N)MTP]. Secondly, to evaluate the effect of a transient cerebral ischaemia in the same task. Both surgical procedures were chosen as they produce a CNS lesion similar to that described in Alzheimer's Disease (AD). Thirdly, the effect of the acetylcholinesterase inhibitor, donepezil (Aricept(R), E2020), on the resulting cognitive impairment was studied. Perforant path transection produced a robust, delay-dependent impairment of choice accuracy in rats performing either a delayed matching- or nonmatching-to-position task. Sample latency was also reduced following lesion, yet the lesion-induced impairment was not affected by increasing the response requirement at the sample stage. An 11-min period of transient ischaemia (two-vessel occlusion model) resulted in almost complete loss of hippocampal CA1 pyramidal cells and a delay-dependent impairment in DMTP performance. However, unlike perforant path lesions, this deficit was unstable and declined in magnitude over the experimental period. Increasing the delay interval restored this deficit. Donepezil, at doses that robustly attenuated a scopolamine (0.06 mg/kg s.c.)-induced DMTP accuracy impairment in naïve, unoperated rats, had no effect against either lesion-induced impairment. The results are considered in terms of the effectiveness of acetylcholinesterase inhibitors in noncholinergic-based preclinical cognitive models.

A novel semi-automated paramagnetic microparticle based enzyme immunoassay for hepatitis C virus: its application to serologic testing.

A new rapid serologic enzyme immunoassay for antibodies to hepatitis C virus (HCV) is described. The assay combines synthetic peptide and recombinant antigens representing putative structural and non structural HCV gene products with paramagnetic microparticle assay (MP assay) technology. Assay readout is based upon an enzymatically generated fluorescent product which is quantified with a novel semi-automated washer/reader instrument system. Assay sensitivity and specificity was determined to be greater than the first generation HCV C-100 EIA using a non-A, non-B hepatitis disease panel, an HCV performance panel, an HCV seroconversion panel, dilutions of HCV reactive sera, and random volunteer blood donor specimens.

A rapid method for purifying PCR products for direct sequence analysis.

An HPLC approach for purification and sequencing of double-stranded DNA obtained directly from a PCR is described. This simple and reliable procedure has several advantages; the DNA fragment is rapidly eluted (less than 7 minutes), requires no organic cleanup, produces several hundred bases of sequence and is sensitive enough to obtain DNA sequence from a single 100-microliters PCR. This method is demonstrated by sequencing tumor necrosis factor alpha (TNF alpha) gene amplified from mouse tail DNA.

Genomic structure and expression of human beta-1,4-galactosyltransferase.

We have cloned 60 kilobases of overlapping human genomic DNA comprising the complete coding sequence of the biosynthetic enzyme beta-1,4-galactosyltransferase (GalTase). The human locus spans greater than 50 kb of genomic DNA and shows an exon structure similar to the mouse gene. However, contrary to the mouse and bovine systems, which yield two distinct transcripts, RNA blotting and RNase protection analysis of human mRNA derived from HeLa cells reveal only a single transcript, corresponding to the "short" form of the mouse and bovine transcripts. Furthermore, Northern analysis on a panel of human cell lines of varying tissue origin and morphology shows GalTase expression levels to be highly variable, consistent with the notion that GalTase expression and consequent cell-specific differences in galactosylation are at least partially regulated at the transcriptional level.