Heterogeneity after harmonisation: A retrospective cohort study of bleeding and stroke risk after the introduction of direct oral anticoagulants in four Western European countries.

PURPOSE: Database heterogeneity can impact effect estimates. Harmonisation provided by common protocols and common data models (CDMs) can increase the validity of pharmacoepidemiologic research. In a case study measuring the changes in the safety and effectiveness of stroke prevention therapy after the introduction of direct oral anticoagulants (DOACs), we performed an international comparison. METHODS: Using data from Stockholm, Denmark, Scotland and Norway, harmonised with a common protocol and CDM, two calendar-based cohorts were created: 2012 and 2017. Patients with a diagnosis code of atrial fibrillation 5 years preceding the 1-year cohort window were included. DOAC, vitamin K antagonist and aspirin treatment were assessed in the 6 months prior to the start of each year while strokes and bleeds were assessed during the year. A Poisson regression generated incidence rate ratios (IRRs) to compare outcomes from 2017 to 2012 adjusted for changes in individual-level baseline characteristics. RESULTS: In 280 359 patients in the 2012 cohort and 356 779 in the 2017 cohort, treatment with OACs increased on average from 45% to 65%, while treatment with aspirin decreased from 30% to 10%. In all countries except Scotland, there were decreases in the risk of stroke and no changes in bleeding risk, after adjustment for changes in baseline characteristics. In Scotland, major bleeding (IRR 1.09, 95% confidence interval [CI] [1.00; 1.18]) and intracranial haemorrhage (IRR 1.31, 95% CI [1.13; 1.52]) increased from 2012 to 2017. CONCLUSIONS: Stroke prevention therapy improved from 2012 to 2017 with a corresponding reduction in stroke risk without increasing the risk of bleeding in all countries, except Scotland. The heterogeneity that remains after methodological harmonisation can be informative of the underlying population and database.

Highly sensitive quantification of optic neuritis intrathecal biomarker CXCL13.

BACKGROUND: Elevation of CXCL13, a key regulator of B-cell recruitment in cerebrospinal fluid (CSF) is implicated in multiple sclerosis (MS). OBJECTIVE: to evaluate if measurement of CXCL13 using a highly sensitive assay is of value in acute optic neuritis (ON) patients for the prediction of later MS. METHOD: CXCL13 was measured by Simoa in two independent treatment-naïve ON cohorts, a training cohort (TC, n = 33) originating from a population-based cohort, a validation cohort (VC, n = 30) consecutively collected following principles for population studies. Prospectively, 14/33 TC and 12/30 VC patients progressed to MS (MS-ON) while 19/33 TC and 18/30 VC patients, remained as isolated ON (ION). RESULTS: CXCL13 was detectable in all samples and were higher in ON compared with healthy controls (HC) (p = 0.012). In the TC, CSF levels in MS-ON were higher compared with ION patients and HC (p = 0.0001 and p<0.0001). In the VC, we confirmed the increase of CXCL13 in MS-ON compared to ION (p = 0.0091). Logistic regression analysis revealed an area under receiver operating characteristic curve of 0.83 [95% C.I: 0.73-0.93]. CONCLUSIONS: The highly sensitive CXCL13 Simoa assay demonstrated ability to identify ON patients and separate MS-ON from ION, and predictive diagnostic values indicates a promising potential of this assay.

CARGEL Bioscaffold improves cartilage repair tissue after bone marrow stimulation in a minipig model.

PURPOSE: To gain knowledge of the repair tissue in critically sized cartilage defects using bone marrow stimulation combined with CARGEL Bioscaffold (CB) compared with bone marrow stimulation (BMS) alone in a validated animal model. METHODS: Six adult Göttingen minipigs received two chondral defects in each knee. The knees were randomized to either BMS combined with CB or BMS alone. The animals were euthanized after 6 months. Follow-up consisted of histomorphometry, immunohistochemistry, semiquantitative scoring of the repair tissue (ICRS II), and µCT of the trabecular bone beneath the defect. RESULTS: There was significantly more fibrocartilage (80% vs 64%, p = 0.04) and a trend towards less fibrous tissue (15% vs 30%, p = 0.05) in the defects treated with CB. Hyaline cartilage was only seen in one defect treated with CB and none treated with BMS alone. For histological semiquantitative score (ICRS II), defects treated with CB scored lower on subchondral bone (69 vs. 44, p = 0.04). No significant differences were seen on the other parameters of the ICRS II. Immunohistochemistry revealed a trend towards more positive staining for collagen type II in the CB group (p = 0.08). µCT demonstrated thicker trabeculae (p = 0.029) and a higher bone material density (p = 0.028) in defects treated with CB. CONCLUSION: Treatment of cartilage injuries with CARGEL Bioscaffold seems to lead to an improved repair tissue and a more pronounced subchondral bone response compared with bone marrow stimulation alone. However, the CARGEL Bioscaffold treatment did not lead to formation of hyaline cartilage.

Cerebrospinal fluid biomarkers for predicting development of multiple sclerosis in acute optic neuritis: a population-based prospective cohort study.

BACKGROUND: Long-term outcome in multiple sclerosis (MS) depends on early treatment. In patients with acute optic neuritis (ON), an early inflammatory event, we investigated markers in cerebrospinal fluid (CSF), which may predict a diagnosis of MS. METHODS: Forty patients with acute ON were recruited in a prospective population-based cohort with median 29 months (range 19-41) of follow-up. Paired CSF and serum samples were taken within 14 days (range 2-38), prior to treatment. Prospectively, 16/40 patients were by a uniform algorithm diagnosed with MS (MS-ON) and 24 patients continued to manifest isolated ON (ION) during follow-up. Levels of cytokines and neurofilament light chain (NF-L) were measured at the onset of acute ON and compared to healthy controls (HC). Significance levels were corrected for multiple comparisons ("q"). The predictive value of biomarkers was determined with multivariable prediction models using nomograms. RESULTS: CSF TNF-α, IL-10, and CXCL13 levels were increased in MS-ON compared to those in ION patients (q = 0.021, 0.004, and 0.0006, respectively). MS-ON patients had increased CSF pleocytosis, IgG indices, and oligoclonal bands (OCBs) compared to ION (q = 0.0007, q = 0.0058, and q = 0.0021, respectively). CSF levels of IL-10, TNF-a, IL-17A, and CXCL13 in MS-ON patients correlated with leukocyte counts (r > 0.69 and p < 0.002) and IgG index (r > 0.55, p < 0.037). CSF NF-L levels were increased in ON patients compared to those in HC (q = 0.0077). In MS-ON, a progressive increase in NF-L levels was observed at 7 to 14 days after disease onset (r = 0.73, p < 0.0065). Receiver-operating characteristic (ROC) curves for two multivariable prediction models were generated, with IL-10, CXCL13, and NF-L in one ("candidate") and IgG index, OCB, and leukocytes in another ("routine"). Area under the curve was 0.89 [95% CI 0.77-1] and 0.86 [0.74-0.98], respectively. Predictions of the risk of MS diagnosis were illustrated by two nomograms. CONCLUSIONS: CSF TNF-α, IL-10, CXCL13, and NF-L levels were associated with the development of MS, suggesting that the inflammatory and neurodegenerative processes occurred early. Based on subsequent diagnosis, we observed a high predictive value of routine and candidate biomarkers in CSF for the development of MS in acute ON. The nomogram predictions may be useful in the diagnostic work-up of MS.

Distinguishing pathogenic mutations from background genetic noise in cardiology: The use of large genome databases for genetic interpretation.

Advances in clinical genetic testing have led to increased insight into the human genome, including how challenging it is to interpret rare genetic variation. In some cases, the ability to detect genetic mutations exceeds the ability to understand their clinical impact, limiting the advantage of these technologies. Obstacles in genomic medicine are many and include: understanding the level of certainty/uncertainty behind pathogenicity determination, the numerous different variant interpretation-guidelines used by clinical laboratories, delivering the certain or uncertain result to the patient, helping patients evaluate medical decisions in light of uncertainty regarding the consequence of the findings. Through publication of large publicly available exome/genome databases, researchers and physicians are now able to highlight dubious variants previously associated with different cardiac traits. Also, continuous efforts through data sharing, international collaborative efforts to develop disease-gene-specific guidelines, and computational analyses using large data, will indubitably assist in better variant interpretation and classification. This article discusses the current, and quickly changing, state of variant interpretation resources within cardiovascular genetic research, e.g., publicly available databases and ways of how cardiovascular genetic counselors and geneticists can aid in improving variant interpretation in cardiology.

Continuing professional development (CPD) in radiography: A collaborative European meta-ethnography literature review.

OBJECTIVES: The aim of the study was to complete a collaborative review of Radiography continuing professional development (CPD) research material to support the production of European Federation of Radiographer Societies (EFRS) CPD recommendations. A meta-ethnography approach to literature review was applied focussing upon commonalities rather than discrepancies between research outcomes. This facilitated exploration of context across the geographical region of Europe with national variations in CPD governance. The seven phases of the meta-ethnographic approach were followed by two independent experienced researchers. A third researcher mediated the findings which were then explored collaboratively with the EFRS CPD working group for concordance. KEY FINDINGS: Phase seven of the meta-ethnography involved interpreting an expression of the synthesis from the previous stages. Six main corroborating themes emerged in this process and following mediation were expressed as themes; knowledge, skills & competency, needs/gap analysis, multi-layered/multi-modal, barriers and drivers; regulation vs autonomy; fostering collaboration - harnessing technology. CONCLUSION: The primary feature of CPD activity should be the resulting impact - to patients, the service, the profession and the individual; with all stakeholders working in partnership. CPD activity must be flexible/multi-modal to support the changing growth/dynamic workforce. All stakeholders should utilise communication and technology resources and make efforts to improve collaboration between the management, regulators and educators to support Radiographers to develop meaningful CPD. Health services across Europe are under increasing stress and a principal factor going forwards will be managing increasing demands on healthcare staff whilst supporting enhancement of the knowledge, skills and competency base.

Hydrochlorothiazide use is strongly associated with risk of lip cancer.

BACKGROUND: The diuretic hydrochlorothiazide is amongst the most frequently prescribed drugs in the United States and Western Europe, but there is suggestive evidence that hydrochlorothiazide use increases the risk of lip cancer. OBJECTIVES: To study the association between use of hydrochlorothiazide and squamous cell carcinoma of the lip. METHODS: We conducted a case-control study using Danish nationwide registry data. From the Cancer Registry (2004-2012), we identified 633 case patients with squamous cell carcinoma (SCC) of the lip and matched them to 63 067 population controls using a risk-set sampling strategy. Hydrochlorothiazide use (1995-2012) was obtained from the Prescription Registry and defined according to cumulative use. Applying conditional logistic regression, we calculated odds ratios (ORs) for SCC lip cancer associated with hydrochlorothiazide use, adjusting for predefined potential confounders obtained from demographic, prescription and patient registries. RESULTS: Ever-use of hydrochlorothiazide was associated with an adjusted OR for SCC lip cancer of 2.1 (95% confidence interval (CI): 1.7-2.6), increasing to 3.9 (95%CI: 3.0-4.9) for high use (≥25 000 mg). There was a clear dose-response effect (P < 0.001), with the highest cumulative dose category of hydrochlorothiazide (≥100 000 mg) presenting an OR of 7.7 (95%CI: 5.7-10.5). No association with lip cancer was seen with use of other diuretics or nondiuretic antihypertensives. Assuming causality, we estimated that 11% of the SCC lip cancer cases could be attributed to hydrochlorothiazide use. CONCLUSIONS: Hydrochlorothiazide use is strongly associated with an increased risk of lip cancer.

Integration of 60,000 exomes and ACMG guidelines question the role of Catecholaminergic Polymorphic Ventricular Tachycardia-associated variants.

Catecholaminergic Polymorphic Ventricular Tachycardia (CPVT) is a highly lethal cardiac arrhythmia disease occurring during exercise or psychological stress. CPVT has an estimated prevalence of 1:10,000 and has mainly been associated with variants in calcium-regulating genes. Identification of potential false-positive pathogenic variants was conducted by searching the Exome Aggregation Consortium (ExAC) database (n = 60,706) for variants reported to be associated with CPVT. The pathogenicity of the interrogated variants was assessed using guidelines from the American College of Medical Genetics and Genomics (ACMG) and in silico prediction tools. Of 246 variants 38 (15%) variants previously associated with CPVT were identified in the ExAC database. We predicted the CPVT prevalence to be 1:132. The ACMG standards classified 29% of ExAC variants as pathogenic or likely pathogenic. The in silico predictions showed a reduced probability of disease-causing effect for the variants identified in the exome database (p < 0.001). We have observed a large overrepresentation of previously CPVT-associated variants in a large exome database. Based on the frequency of CPVT in the general population, it is less likely that the previously proposed variants are associated with a highly penetrant monogenic form of the disease.

Resistin Gene Expression is Downregulated in CD4(+) T Helper Lymphocytes and CD14(+) Monocytes in Rheumatoid Arthritis Responding to TNF-α Inhibition.

Rheumatoid arthritis (RA) is caused by complex interactions between immune cells and sustained by Th1 response cytokines. Resistin [resistance to insulin; (RETN)] is an inflammatory cytokine, first discovered in murine adipocytes. In man, RETN is mainly secreted by monocytes. The distinct role of RETN in the immune reaction is uncertain; however, RETN has pro-inflammatory, pro-fibrotic and possibly tolerogenic properties. The aim was to assess the reaction of RETN gene expression to TNF-α inhibition (I) in pathogenetic immune cell subsets in RA, in the context of Th1, inflammatory and regulatory cytokine gene expressions. Accordingly, we measured RETN, IFN-γ, TNF-ß, IL-1ß, TNF-α, TGF-ß and IL-10 gene expressions in CD14(+) monocytes, CD4(+) T helper (Th) lymphocytes (ly), CD8(+) T cytotoxic (Tc) ly and CD19(+) B ly in active RA before and 3 months after start of TNF-αI. Leucocyte subsets were separated by specific monoclonal antibody-covered beads, RNA extracted and levels of RETN, Th1 response, inflammatory and regulatory cytokine mRNAs measured by quantitative reverse transcription-polymerase chain reaction technique. We found that TNF-αI caused a significant downregulation of RETN gene expression in CD14(+) monocytes and CD4(+) Th ly and was unchanged in CD8(+) Tc ly and CD19(+) B ly. Both in active RA and during TNF-αI, RETN mRNA levels were significantly higher in CD14(+) monocytes than in all other examined cell types. In monocytes, fold change in RETN and TGF-ß gene expressions upon TNF-αI correlated significantly. Our findings indicate that RETN has pro-inflammatory as well as proresolving roles in active RA.

Mucin-mediated nanocarrier disassembly for triggered uptake of oligonucleotides as a delivery strategy for the potential treatment of mucosal tumours.

This work demonstrates gastric mucin-triggered nanocarrier disassembly for release of antisense oligonucleotides and consequent unassisted cellular entry as a novel oral delivery strategy. A fluorescence activation-based reporter system was used to investigate the interaction and mucin-mediated disassembly of chitosan-based nanocarriers containing a 13-mer DNA oligonucleotide with a flanked locked RNA nucleic acid gapmer design. Gastric mucins were shown to trigger gapmer release from nanocarriers that was dependent on the interaction time, mucin concentration and N : P ratio with a maximal release at N : P 10. In contrast to siRNA, naked gapmers exhibited uptake into mucus producing HT-MTX mono-cultures and HT-MTX co-cultured with the carcinoma epithelial cell line Caco-2. Importantly, in vivo gapmer uptake was observed in epithelial tissue 30 min post-injection in murine intestinal loops. The findings present a mucosal design-based system tailored for local delivery of oligonucleotides that may maximize the effectiveness of gene silencing therapeutics within tumours at mucosal sites.

Anxiolytic-like effects after vector-mediated overexpression of neuropeptide Y in the amygdala and hippocampus of mice.

Neuropeptide Y (NPY) causes anxiolytic- and antidepressant-like effects after central administration in rodents. These effects could theoretically be utilized in future gene therapy for anxiety and depression using viral vectors for induction of overexpression of NPY in specific brain regions. Using a recombinant adeno-associated viral (rAAV) vector, we addressed this idea by testing effects on anxiolytic- and depression-like behaviours in adult mice after overexpression of NPY transgene in the amygdala and/or hippocampus, two brain regions implicated in emotional behaviours. In the amygdala, injections of rAAV-NPY caused significant anxiolytic-like effect in the open field, elevated plus maze, and light-dark transition tests. In the hippocampus, rAAV-NPY treatment was associated with anxiolytic-like effect only in the elevated plus maze. No additive effect was observed after combined rAAV-NPY injection into both the amygdala and hippocampus where anxiolytic-like effect was found in the elevated plus maze and light-dark transition tests. Antidepressant-like effects were not detected in any of the rAAV-NPY injected groups. Immobility was even increased in the tail suspension and forced swim tests after intra-amygdaloid rAAV-NPY. Taken together, the present data show that rAAV-NPY treatment may confer non-additive anxiolytic-like effect after injection into the amygdala or hippocampus, being most pronounced in the amygdala.

High prevalence of genetic variants previously associated with Brugada syndrome in new exome data.

More than 300 variants in 12 genes have been associated with Brugada syndrome (BrS) which has a prevalence ranging between 1:2000 and 1:100,000. Until recently, there has been little knowledge regarding the distribution of genetic variations in the general population. This problem was partly solved, when exome data from the NHLI GO Exome Sequencing Project (ESP) was published. In this study, we aimed to report the prevalence of previously BrS-associated variants in the ESP population. We performed a search in ESP for variants previously associated with BrS. In addition, four variants in ESP were genotyped in a second Danish control population (n = 536) with available electrocardiograms. In ESP, we identified 38 of 355 (10%) variants, distributed on 272 heterozygote carriers and two homozygote carriers. The genes investigated were on average screened in 6258 individuals. This corresponds to a surprisingly high genotype prevalence of 1:23 (274:6258). Genotyping the four common ESP-derived variants CACNA2D1 S709N, SCN5A F2004L, CACNB2 S143F, and CACNB2 T450I in the Danish controls, we found a genotype prevalence comparable with that found in ESP. We suggest that exome data are used in research, as an additive tool to predict the pathogenicity of variants in patients suspected for BrS.

The primary Maxillary Central Incisor in the Solitary Median Maxillary Central Incisor syndrome.

AIM: Solitary Median Maxillary Central Incisor (SMMCI) is a developmental anomaly in the permanent dentition with one single central incisor in the maxilla, positioned exactly in the midline. This condition has been associated with extra- and intraoral malformations in the frontonasal segment of the cranium and face. It is not known whether the centrally located permanent incisor is always preceded by a centrally located primary incisor. The aim was to analyse whether a permanent single central incisor in SMMCI is always preceded by a primary single central incisor and to study extra- and intraoral phenotypic traits of the condition. STUDY DESIGN: cross-sectional radiographic study of 11 children, visual analysis of photos and dental and panoramic radiographs. RESULTS: Nine of the 11 cases exhibited a primary SMMCI with one symmetrical crown and root. Two cases exhibited two separate primary central incisor crowns with fused roots. The phenotypical traits (indistinct philtrum, lack of normal upper lip contour, missing superior labial frenulum and distinct mid-palatal ridge) were findings observed in young children with a primary SMMCI. CONCLUSION: The present study concludes and stresses the necessity of diagnosing of the SMMCI condition early in life. Furthermore, paediatric dentists are recommended to be aware of the condition and to refer these patients to interdisciplinary diagnostics and treatment.

Y5 neuropeptide Y receptor overexpression in mice neither affects anxiety- and depression-like behaviours nor seizures but confers moderate hyperactivity.

Neuropeptide Y (NPY) has been implicated in anxiolytic- and antidepressant-like behaviour as well as seizure-suppressant effects in rodents. Although these effects appear to be predominantly mediated via other NPY receptors (Y1 and/or Y2), several studies have also indicated a role for Y5 receptors. Gene therapy using recombinant viral vectors to induce overexpression of NPY, Y1 or Y2 receptors in the hippocampus or amygdala has previously been shown to modulate emotional behaviour and seizures in rodents. The present study explored the potential effects of gene therapy with the Y5 receptor, by testing effects of recombinant adeno-associated viral vector (rAAV) encoding Y5 (rAAV-Y5) in anxiety- and depression-like behaviour as well as in kainate-induced seizures in adult mice. The rAAV-Y5 vector injected into the hippocampus and amygdala induced a pronounced and sustained increase in Y5 receptor mRNA expression and functional Y5 receptor binding, but no significant effects were found with regard to anxiety- and depression-like behaviours or seizure susceptibility. Instead, rAAV-mediated Y5 receptor transgene overexpression resulted in moderate hyperactivity in the open field test. These results do not support a potential role for single transgene overexpression of Y5 receptors for modulating anxiety-/depression-like behaviours or seizures in adult mice. Whether the induction of hyperactivity by rAAV-Y5 could be relevant for other conditions remains to be studied.

Neuropeptide Y Y1 receptor hippocampal overexpression via viral vectors is associated with modest anxiolytic-like and proconvulsant effects in mice.

Neuropeptide Y (NPY) exerts anxiolytic- and antidepressant-like effects in rodents that appear to be mediated via Y1 receptors. Gene therapy using recombinant viral vectors to induce overexpression of NPY in the hippocampus or amygdala has previously been shown to confer anxiolytic-like effect in rodents. The present study explored an alternative and more specific approach: overexpression of Y1 receptors. Using a recombinant adeno-associated viral vector (rAAV) encoding the Y1 gene (rAAV-Y1), we, for the first time, induced overexpression of functional transgene Y1 receptors in the hippocampus of adult mice and tested the animals in anxiety- and depression-like behavior. Hippocampal Y1 receptors have been suggested to mediate seizure-promoting effect, so the effects of rAAV-induced Y1 receptor overexpression were also tested in kainate-induced seizures. Y1 receptor transgene overexpression was found to be associated with modest anxiolytic-like effect in the open field and elevated plus maze tests, but no effect was seen on depression-like behavior using the tail suspension and forced swim tests. However, the rAAV-Y1 vector modestly aggravated kainate-induced seizures. These data indicate that rAAV-induced overexpression of Y1 receptors in the hippocampus could confer anxiolytic-like effect accompanied by a moderate proconvulsant adverse effect. Further studies are clearly needed to determine whether Y1 gene therapy might have a future role in the treatment of anxiety disorders.

Fluoxetine reverts chronic restraint stress-induced depression-like behaviour and increases neuropeptide Y and galanin expression in mice.

Stressful life events and chronic stress are implicated in the development of depressive disorder in humans. Neuropeptide Y (NPY) and galanin have been shown to modulate the stress response, and exert antidepressant-like effects in rodents. To further investigate these neuropeptides in depression-like behaviour, NPY and galanin gene expression was studied in brains of mice subjected to chronic restraint stress (CRS) and concomitant treatment with the antidepressant fluoxetine (FLX). CRS caused a significant increase in depression-like behaviour that was associated with increased NPY mRNA levels in the medial amygdala. Concomitant FLX treatment reverted depression-like effects of CRS and led to significant increases in levels of NPY and galanin mRNA in the dentate gyrus, amygdala, and piriform cortex. These findings suggest that effects on NPY and galanin gene expression could play a role in the antidepressant effects of FLX.

Efficacy of partially hydrolyzed guar gum-added oral rehydration solution in the treatment of severe cholera in adults.

BACKGROUND: Partially hydrolyzed guar gum (PHGG) is a water-soluble fiber if added to oral rehydration solution (ORS) and undergoes fermentation in the colon liberating short chain fatty acids (SCFAs). SCFAs potentiate the effect of ORS, reducing the severity of diarrhea. AIM: To examine the effect of PHGG-added ORS in reducing the stool output and duration of diarrhea in adult cholera. METHODS: 195 male patients were studied in a randomized controlled trial: (a) 65 received ORS + 25 g PHGG; (b) 65 received ORS + 50 g PHGG, and (c) 65 received ORS alone (control). Major outcomes were stool weight and duration of diarrhea. RESULTS: No significant differences were found in mean +/- SD stool weight (g/kg b.w.) during the first and second 24 h. In the subgroup analysis (excluding very high purging patients, stool weight in the first 24 h was >10 kg), the stool weight (g/kg b.w.) was significantly reduced in the first 24 h in both groups receiving PHGG (PHGG 25 g, 136 +/- 68 vs. PHGG 50 g, 144 +/- 49 vs. control, 176 +/- 43, p = 0.01). CONCLUSION: PHGG-added ORS might have a beneficial effect in moderately purging adult cholera. However, further studies are warranted to confirm the preliminary findings.

Unaltered neuropeptide Y (NPY)-stimulated [35S]GTPgammaS binding suggests a net increase in NPY signalling after repeated electroconvulsive seizures in mice.

Although electroconvulsive seizures (ECS) are widely used as a treatment for severe depression, the working mechanism of ECS remains unclear. Repeated ECS causes anticonvulsant effects that have been proposed to underlie the therapeutic effect of ECS, and neuropeptide Y (NPY) is a potential candidate for mediating this anticonvulsant effect. Repeated ECS results in prominent increases in NPY synthesis. In contrast, NPY-sensitive receptor binding is decreased, so it is unclear whether ECS causes a net increase in NPY signalling. Agonist-stimulated [35S]GTPgammaS binding is a method for detecting functional activation of G-protein-coupled receptors. The present study in mice examined the effects of daily ECS for 14 days on NPY-stimulated [35S]GTPgammaS functional binding and compared this with gene expression of NPY and NPY receptors as well as [125I]peptide YY (PYY) binding in hippocampus of the same animals. Significant increases in NPY mRNA and concomitant reductions in NPY-sensitive binding were found in the dentate gyrus, hippocampal CA1, and neocortex of ECS treated mice, which is consistent with previous rat data. These changes remained significant 1 week after repeated ECS. Significant increases in NPY Y1, Y2, and Y5 mRNA were found in the dentate gyrus after ECS. Surprisingly, unaltered levels of functional NPY receptor binding accompanied the decreased NPY-sensitive binding. This suggests that mechanisms coupling NPY receptor stimulation to G-protein activation could be augmented after repeated ECS. Thus increased synthesis of NPY after repeated ECS should result in a net increase in NPY signalling in spite of reduced levels of NPY-sensitive binding.

Regulation of activity-regulated cytoskeleton protein (Arc) mRNA after acute and chronic electroconvulsive stimulation in the rat.

The temporal profile of Arc gene expression after acute and chronic electroconvulsive stimulations (ECS) was studied using semi-quantitative in situ hybridisation in the rat cortex. A single ECS strongly and temporarily increased Arc mRNA levels in dentate granular cells with maximal induction seen up to 4 h after the stimulus, but returned to baseline at 24 h. A single ECS also increased expression of Arc mRNA in the CA1 and the parietal cortex, but the expression peaked within 1 h and returned to baseline levels within 2 h. Repeated or chronic ECS is a model of electroconvulsive therapy and it would be predicted that gene products involved in antidepressant effects accumulate after repeated ECS. However, repeated ECS reduced Arc gene expression in the CA1 24 h after the last stimulus. These results indicate that Arc is an immediate early gene product regulated by an acute excitatory stimulus, but not accumulated by long term repetitive ECS and therefore not a molecular biomarker for antidepressant properties. More likely, Arc is likely a molecular link to the decline in memory consolidation seen in depressive patients subjected to electroconvulsive therapy.