Amyloid fibrils are rich in β-sheet and can be observed with thioflavin

Natural Product Library T (ThT) assay or by staining with Congo red, indicating that they contain a hydrophobic region. Although these fibrillar amyloids were previously considered to be the primary factor in the induction of pathology in these protein conformational diseases, recent studies indicate that small oligomers or protofibrils, rather than amyloid fibrils, may play an important role in cytotoxicity (Lesnéet al., 2006; Haataja et al., 2008). In this study, we compared TDH and TRH to investigate whether membrane toxicity by the toxins is induced by amyloidogenicity upon heating or small oligomerized tetrameric structures. TRH showed less amyloidogenicity compared with that of TDH. However, the hemolytic activity of TRH was similar to that of TDH. These data indicate that membrane disruption by the TDH family is mediated by tetrameric structures and not by the amyloidogenicity. We also compared

the circular dichroism (CD) spectra of TDH and TRH in the heat-denatured state and found that an incorrect Omipalisib purchase refolding process resulted in loss of the Arrhenius effect of TRH. Purification of recombinant TDH was performed as described previously (Naim et al., 2001). N-terminal signal peptide-deleted (1–24 amino acids) trh1 (GenBank accession no. AB112353) was inserted into the expression vector pET-28a (Novagen). For the expression of recombinant TRH, we transformed a plasmid vector pET28-a harboring trh1 gene into Escherichia coli JM109 (DE3) cells (Promega). The transformant was cultured in Luria–Bertani broth (1% Bacto tryptone, 0.5% yeast extract, and 1% NaCl) containing 100 μg mL−1 of kanamycin at 30 °C for 30 h with rotary shaking, and then centrifuged at 6000 g for 30 min. We added ammonium sulfate (55% saturation) to the supernatant and allowed it to stir overnight, followed by centrifugation at 10 000 g for 1 h. The pellet was Cyclin-dependent kinase 3 dissolved in 10 mM phosphate buffer

(pH 7.4) and dialyzed against the same buffer. We applied this solution to a series of columns: Cellulofine Hap (hydroxyapatite) (Seikagaku-Kogyo, Tokyo, Japan), Toyopearl DEAE-650M (Tosoh, Tokyo, Japan), Resource-Phe (Amersham Pharmacia Biotech AB, Uppsala, Sweden), and Superose 6 (GE Healthcare, Uppsala, Sweden). Hemolytic activities were measured as described previously (Fukui et al., 2005). Far-UV CD spectra were recorded with a J-720W spectropolarimeter (Jasco, Tokyo, Japan) equipped with a thermoelectric temperature controller. Data were analyzed with the software provided by Jasco. Measurements were taken in a quartz cuvette with a path length of 2 mm, scanned in steps of 0.2 nm at a rate of 50 nm min−1. Samples of 0.2 mg mL−1 TRH in 10 mM phosphate buffer (pH 7.4) were heated up from 37 to 90 °C at a heating rate of 0.1 °C min−1. After heat treatment at 90 °C, the temperature was decreased rapidly by 30 °C min−1 or slowly by 1 °C min−1 decrements to 37 °C.

Also, it is possible that patients may have a store of drugs not previously used. By its nature, this adherence measure does not provide any

information about short-term adherence patterns and whether the FK228 in vivo antiretroviral drugs were taken at the correct times. However, a gold standard for measuring adherence does not exist [47]. A feature that makes adherence particularly difficult to accurately quantify is that rates may vary not just between individuals, but also within the same individual over time. This is the reason why we decided to evaluate drug coverage on the last 6 months immediately previous to time-zero, at every DCVL episode rather than assess it only in one point over time. The second limitation is that the risk of viral rebound may be overestimated, as a result of the definition of VL rebound as a single VL >200 copies/mL. Thus, it represents a relatively low threshold and is not necessarily confirmed by a subsequent VL >200 copies/mL, but our aim selleck chemicals llc was to detect even transient increase of plasma VL. It should be noted that patients may spontaneously re-suppress the VL after a single episode of viraemia >200 copies/mL. Finally, because this study included only subjects who had achieved

viral suppression, the results regarding the relationship between adherence and outcome can only be generalized to HIV-infected subjects on HAART who achieved VL suppression, who represent approximately 90% of HIV-infected patients on HAART [48]. To conclude, although it is well established that most patients on ART nowadays achieve VL suppression, full and long-term maintenance of this suppression, which is the current accepted goal of therapy [49], can be problematic [16]. In this study we have confirmed the importance of ART adherence, as evaluated by drug coverage, to predict VL rebound. Therefore, objective, simple and easy-to-use adherence measures, such as

the one proposed here, could help to identify, together with other known predictors, O-methylated flavonoid patients at risk of viral rebound, thereby guiding corrective interventions to prevent and promptly manage viral rebound. This work was funded in part by NEAT (European Commission). Royal Free Centre for HIV Medicine Clinical: S. Bhagani, P. Byrne, A. Carroll, I. Cropley, Z. Cuthbertson, A. Dunleavy, A. M. Geretti, B. Heelan, M. Johnson, S. Kinloch-de Loes, M. Lipman, S. Madge, T. Mahungu, N. Marshall, D. Nair, B. Prinz, A. Rodger, L. Swaden, M. Tyrer and M. Youle. Data management: C. Chaloner, H. Grabowska, J. Holloway, J. Puradiredja, S. Scott and R. Tsintas. Biostatistics/Epidemiology: W. Bannister, L. Bansi, V. Cambiano, A. Cozzi-Lepri, Z. Fox, E. Harris, T. Hill, A. Kamara, F. Lampe, R. Lodwick, A. Mocroft, A. Phillips, J. Reekie, A. Rodger, C. Sabin and C. Smith. Laboratory: E. Amoah, C. Booth, G. Clewley, A. Garcia Diaz, A. M. Geretti, B.

In order to avoid disruption of the clinic’s flow, brief post-test counselling could be provided after the patient receives the preliminary result and more extensive counselling planned for the next consultation

when the patient MAPK Inhibitor Library order returns for the confirmation result. Robust links between primary care and HIV specialist services are essential for immediate linkage to care for newly diagnosed patients in order to minimize loss to follow-up. Targeting testing at patients considered to be at high risk of HIV infection in line with World Health Organization (WHO) and the European Centre for Disease Prevention and Control (ECDC) recommendations, as proposed by a majority of the study participants, would make selleck compound implementation cost-effective by reducing the number of tests required. The majority of participating GPs were aware of the existence of rapid HIV tests but did not know how to use them. Specific training in rapid HIV testing should be offered to Spanish GPs. Efforts have to be made to improve training in the provision of pre-test information

and post-test counselling and to improve skills in sexual history taking, in order to identify those patients at risk. Also, GPs should be made aware of missed opportunities for HIV testing and the necessity of their involvement in the early diagnosis of HIV infection. The principal limitations of this study are the opportunistic sampling design of the survey, making the results difficult to generalize to all Spanish GPs, and the low return rate for questionnaires. This response rate is, however, similar to that seen in other surveys of similar study populations and the sample size achieved is greater than in most comparable studies. It is also likely that the GPs who responded are those with a greater interest in HIV/AIDS and hence those most likely to take on board any changes in testing policy likely to have an impact on testing rates. In summary, early

HIV diagnosis and timely linkage to care should be one of the main strategies to both improve the prognosis of HIV-positive BCKDHB patients and decrease the incidence of HIV infection in the community. In most settings, primary health care is a frontline service for people with symptoms of acute infection or at risk of HIV infection and other sexually transmitted infections. Our data demonstrate the openness of these professionals to introducing rapid HIV testing into primary health care and moreover identify the main barriers to doing so. According to our results, the introduction of rapid test technology in the primary health sector may be useful in increasing the number of test performed at this level.

Natural and recombinant Alt a 1 proteins share secondary structure and IgE-binding determinants and skin testing shows a similar reactivity. These results confirm that rAlt a 1 could be an effective candidate for the development of diagnostic and therapeutic approaches and that Y. lipolytica has become an attractive host for the expression of complex proteins such allergens. The authors thank Dr A. R. Viguera (Unidad de Biofísica, Universidad del País Vasco-CSIC, Leioa, Spain) for CD spectra analysis. J.A.A. is employee Rucaparib cell line of the biopharmaceutical company Bial-Arístegui. “
“Isoprenoids are a large, diverse group of secondary metabolites which has recently raised a renewed research

interest due to genetic engineering advances, allowing specific selleck isoprenoids to be produced and characterized in heterologous hosts. Many researches on metabolic engineering of heterologous hosts for increased isoprenoid production are focussed on Escherichia coli and yeasts. E. coli, as most prokaryotes, use the 2-C-methyl-d-erythritol-4-phosphate (MEP) pathway for isoprenoid production. Yeasts on the other hand, use the mevalonate pathway which is commonly found in eukaryotes. However, Lactococcus lactis is an attractive alternative

host for heterologous isoprenoid production. Apart from being food-grade, this Gram-positive prokaryote uses the mevalonate pathway for isoprenoid production instead of the MEP pathway. Previous studies have shown that L. lactis is able to produce sesquiterpenes through heterologous expression of plant sesquiterpene synthases. In this work, we analysed the gene expression of the lactococcal mevalonate pathway through RT-qPCR to successfully engineer L. lactis as an efficient host for isoprenoid production. We then overexpressed the mvk gene singly or co-expressed with the mvaA gene as an attempt Pregnenolone to increase β-sesquiphellandrene production in L. lactis. It was observed that co-expression of mvk with mvaA doubled the amount of β-sesquiphellandrene produced. “
“Myxopyronin B (MyxB) binds to the switch region

of RNA polymerase (RNAP) and inhibits transcriptional initiation. To evaluate the potential development of MyxB as a novel class of antibiotic, we characterized the antimicrobial activity of MyxB against Staphylococcus aureus. Spontaneous MyxB resistance in S. aureus occurred at a frequency of 8 × 10−8, similar to that of rifampin. The MyxB-resistant mutants were found to be altered in single amino acid residues in the RNAP subunits that form the MyxB-binding site. In the presence of human serum albumin, the MyxB minimum inhibitory concentration against S. aureus increased drastically (≥128-fold) and 99.5% of MyxB was protein bound. Because of the high serum protein binding and resistance rate, we conclude that MyxB is not a viable starting point for antibiotic development.

Patients were treated according to the clinical picture and the treating physician. We followed up patients until the resolution of symptoms. Fifteen cases of travel-related leptospirosis were included in this study. Nearly all patients (14/15) were men, mean age was 34 years [interquartile range (IQR): 28–52] (Table

2). All travelers except one were tourists. The infection was contracted primarily in Asia (47%). The mean duration of travel was 18 days (IQR: 15–32). The most frequent at-risk exposure was bathing in fresh water (10/15), followed by nautical sporting activities (kayaking, rafting, canoeing) in four cases. We found a history of skin wound in 3 of the 10 patients who had fresh-water exposure. In one patient who had stayed in a rural area, exposure was not established. Four patients, including one expatriate, developed symptoms before their return to France. In the 11 remaining patients, the average lag time Doxorubicin nmr between return to country of origin and onset of symptoms

was 5 days (IQR: 2–7). Fever (temperature >38°C) was found in all patients. Other signs and symptoms selleck included were headache (80%), digestive disorders (67%) (nausea and/or vomiting, diarrhea), myalgias (53%), and arthralgias (47%). Exanthema, jaundice, and hepatosplenomegaly were observed in 20% of patients. Conjunctival suffusion, macroscopic hematuria, and hemoptysis were rarely observed (7%). Table 3 shows laboratory results. Elevation of LFTs was present in 100% of patients [average values: ASAT = 93 IU/L (3N), ALAT = 137 IU/L (4N)]. The majority

of patients had thrombocytopenia (average thrombocyte levels: 93,809/L), lymphocytopenia (average value: lymphocytes = 694/L) together with moderate renal impairment (average value: creatinine = 193 µmol/L (2N). Antibodies to specific Dynein serovars were identified in 13 cases out of 15 (87%): Leptospira sejroe serovar Hardjobovis (n = 3; 1/400, 1/1600, 1/400), Leptospira cynopteri (n = 1; 1/800), Leptospira bataviae (n = 1; 1/1,600), Leptospira grippotyphosa (n = 2; 1/400,1/6,400), Leptospira hebdo (n = 1; 1/200), Leptospira javanica (n = 1; 1/800), Leptospira icterohaemorrhagiae (n = 1; 1/200), Leptospira tarrassovi (n = 2; 1/1,600, 1/6,400), Leptospira canicola (n = 1; 1/800). Hospitalization was required for eight patients (53%). Seven patients (47%) were treated with amoxicillin (1–2 g, three to four times per day for 7–15 days), including four treated with amoxicillin alone, two with amoxicillin and ceftriaxone (because of meningitis), and one with amoxicillin plus spiramycin (because of pneumonia). The other seven treated patients were given doxycycline (n = 4), 200 mg/day for 10 days, ceftriaxone (n = 2) 1 g/day for 10 days or a combination of ceftriaxone, doxycycline, and spiramycin because of severe disease with acute renal failure and pulmonary involvement. One patient was not treated due to delayed diagnosis (6 months).

Written informed consent was obtained from all patients. The study protocol was approved by the appropriate committees and authorities and was conducted in accordance with the Declaration high throughput screening compounds of Helsinki. The cut-off date for this analysis was when all patients had reached week

192 or had discontinued earlier. Efficacy and safety variables were assessed at screening, at baseline, at 2 weeks, at 4 weeks, then every 4 weeks until week 16, at week 24, and every 12 weeks until week 192 (or earlier discontinuation). The primary population for the efficacy analyses was the ITT population. This was used to test for noninferiority with the TLOVR algorithm being followed to assess virological response (HIV-1 RNA < 50 copies/mL). Response RG7422 supplier and loss of response needed to be confirmed and were defined as response/lack of response at two consecutive visits. Intermittent missing values were imputed as responders only if the patient was responding at the preceding and following visits. If noninferiority was established in the study, superiority testing was also to be performed on this population. Efficacy analyses were also performed on the per-protocol (PP) population: all patients who were randomized, who had taken trial medication and who did not take any disallowed antiretroviral medication

as described in the protocol for > 1 week. Noninferiority at week 192 was confirmed if the lower limit of the 95% confidence interval (CI) of the difference between the DRV/r and LPV/r arms was higher than –12%. Superiority

was confirmed if the lower limit of the 95% CI for the difference in treatment response between the treatment arms was greater than 0%. In this study, secondary efficacy variables Oxymatrine included the percentage of patients with plasma viral load < 400 copies/mL at all time-points. Additional sensitivity analyses were also performed to compare virological response rates. These included: PP-TLOVR; ITT missing=failure (M = F); ITT-TLOVR non-VF-censored (patients are censored out after they discontinued for reasons other than VF); and Food and Drug Administration snapshot, whereby only the last HIV-1 RNA measurement within a window of the assessed time-point was taken into account to determine response. Virological response rates were also analysed by the following subgroups: baseline HIV-1 RNA, CD4 cell count, gender, race, age and viral clade. Statistical analyses were carried out using a logistic regression model adjusted for treatment and stratification factors. Changes in CD4 cell count were analysed using the noncompleter=failure (NC = F) imputation. A modified medication adherence self-report inventory (M-MASRI) questionnaire was used to evaluate patient adherence to treatment up to week 192 or time of withdrawal.

Curiously, the stop codons of the convergently oriented ORFs Smlt0783–Smlt0784 and Smlt4197–Smlt4198, are contributed by interleaved

SMAG dimers. The same holds for ORFs Smlt1380–Smlt1381 and Smlt0188–Smlt0189, the stop codons of each being contributed by interleaved Selleck Thiazovivin SMAG trimers. Some SMAGs located between convergently oriented ORFs are at a close distance from the stop codons of both. Accordingly, the number of the ORFs immediately flanked by SMAGs is higher than the number of repeats (501 vs. 406). By contrast, we found only 81 SMAGs located 1–50 bp from ORF stop codons, and 16 that overlap ORF start codons and encode 4–29 aminoacids. About 1/3 of the ORFs flanked 5′ by SMAGs (26/97) carries SMAG sequences also at the 3′ end. K279a ORFs at a close distance from SMAGs are listed in Table S2. Thirty SMAGs are entirely located within ORFs. These repeats can be sorted

into two main groups. Sixteen out of 30 lie within ORFs encoding small hypothetical proteins that do not exhibit significant homology to ORFs encoded by either the S. maltophilia R551-3 or other prokaryotic genomes, and thus plausibly do not correspond to authentic gene products. Similar conclusions were reached for short ORFs interrupted by REPs in Pseudomonas syringae (Tobes & Pareja, 2005). The remaining 14 repeats are found at the same relative genome coordinates in the R551-3 DNA. However, only six interrupted ORFs are conserved in the two strains. SMAGs within ORFs are listed in Table S3. On the whole, intergenic SMAGs are selleckchem found at 747 loci. Of these, 370 separate unidirectionally transcribed ORFs, 343 convergently transcribed ORFs and only 34 divergently transcribed ORFs. The size of repeated DNA families may vary among isolates. To gain a rough estimate of the size of SMAG families scattered in the other two sequenced S. maltophilia genomes, repeats perfectly matching the 40 SMAG sequence variants found in K279a DNA O-methylated flavonoid were searched in R551-3 and SKA14 DNAs. The relative abundance of the five SMAG subfamilies is comparable

in the three genomes. However, their sizes varied, SMAG-2 elements being more abundant in R551-3 and SKA14 and SMAG-3 being predominant in K279a DNA (Fig. 4). The degree of conservation of SMAG sequences was checked by direct sequence comparisons. Thirty-two regions of the K279a chromosome containing SMAG-3 dimers were analyzed in R551-3. Dimers were conserved in 10 regions, missing in nine and replaced in 13 by SMAG-1 or SMAG-2 sequences (monomers or dimers). Fifty K279a intergenic regions containing SMAG-1 HH dimers were also checked in R551-3 DNA. Most (91%) of the K279a SMAG-1 fit the consensus WGCCGGCCgctGGCCGCCW, and have been called α units, and only 4% fit the consensus CGCCGGGCcatGCCCGGCG, and have been called β units (lowercase letters denote loop sequences).

J.M.S.-R. was supported by grant of Consejo Superior de Investigaciones Científicas, CSIC (JAEPre 09 01804). Dr Phillip Wharton is acknowledged for reviewing the English. “
“The Writing Group thanks Dr David Hawkins, Dr Fiona Lyons and Dr Danielle

Mercey for their peer-review of the Guidelines. Dr A de Ruiter has received lecture and Selleck Hydroxychloroquine consultancy fees from Bristol-Myers Squibb and Gilead. Dr GP Taylor’s department has received research grants from Abbott. Dr A Palfreeman has received conference support from Bristol-Myers Squibb and Gilead. Miss P Clayden has no conflicts of interest to declare. Dr J Dhar has received conference support from ViiV. Mrs K Gandhi has no conflicts of interest to declare. Dr Y Gilleece has received lecture and consultancy fees from ViiV. Dr K Harding has no conflicts of interest to declare.

Dr D Hawkins has received lecture fees from Janssen, consultancy fees from Bristol-Myers Squibb, and his department has received research grant support from Bristol-Myers Squibb. Dr P Hay has received lecture and consultancy fees from Abbott, Boehringer Ingelheim, Bristol-Myers Squibb, Gilead, Johnson and Johnson (Tibotec) and ViiV. He has received conference support from Bristol-Myers Y-27632 supplier Squibb, Gilead and Janssen and his department has received research grant support from Abbott, Boehringer Ingelheim, Gilead, Janssen and ViiV. Ms J Kennedy has no conflicts of interest to declare. Dr N Low-Beer has no conflicts of interest to declare. Dr H Lyall has received lecture fees from Danone

and ViiV. Dr F Lyons has no conflicts of interest to declare. Dr D Mercey has no conflicts of interest to declare. Dr P Tookey has no conflicts of interest to declare. Dr S Welch has no conflicts of interest to declare. Dr E Wilkins has received lecture and consultancy fees from Abbott, Bristol-Myers Squibb, Gilead, Janssen, Merck Sharp and Dohme and Pfizer. “
“To study the mechanism of action of the lactobacilli, splenocytes were incubated with lactobacilli. We compared the ability of live and lyophilized Lactobacillus casei, Lactobacillus rhamnosus GG and Lactobacillus delbrueckii ssp. bulgaricus Bortezomib chemical structure to modulate the production of interleukin 12p40 (IL-12p40), tumor necrosis factor α and IL-10 by splenocytes from C57BL/6 and BALB/c mice. Blocking contact between lactobacilli and immune cells abrogated all cytokine production. Toll-like receptor 2 (TLR2) was partially responsible, but not TLR4 or TLR9, for the induction of cytokine production in splenocytes. All cytokine production declined to basal levels when bacterial phagocytosis was inhibited. This shows that lactobacilli stimulation of cytokine production in splenocytes requires the process of phagocytosis and engagement of TLR2, but not TLR4 or TLR9.

However, inherent in this thesis is the notion that greater differential activity should be driven by increased alpha-band suppressive mechanisms during switch trials, i.e. greater synchronisation over http://www.selleckchem.com/products/GDC-0980-RG7422.html frontoparietal control regions. This, however, is not what was found here. Instead, when we made within-modality comparisons of switch vs. repeat trials, a wholly different picture emerged. The increases in differential between-modalities effects were actually driven by greater desynchronisations rather than the predicted increases in synchronisation. Further, these differential effects were entirely driven by changes in alpha-band

power during anticipations of the visual task rather than the auditory task. When switch and repeat trials in anticipation of the auditory task were compared there were essentially no differences found, with late increases in synchronisation of alpha-band activity found to be just as prominent during repeat trials as they were during switch trials. In contrast, desynchronisations of alpha during visual trials were found to be substantially stronger and earlier on switch trials than they were on repeat trials. These more vigorous desynchronisations also showed a more widespread scalp topography that EPZ015666 in vitro included a prominent focus over frontocentral scalp in addition to the more typical parieto-occipital foci. How then do the current results accord

with our original hypothesis? The pattern of behavioral results is instructive here. First, when one compares task performance on mixed-task blocks Montelukast Sodium to that on pure-task blocks, it is clear that the need to switch between tasks had a major impact on task accuracy. Participants were considerably less able to discriminate targets (even on repeat trials) during the blocks in which switching was required as opposed to blocks in which only one task was performed alone over extended periods. On the other hand, the use of instructional pre-cues to indicate which task was to be engaged

during mixed blocks led to the complete alleviation of the classical switch costs that are typically seen during mixed blocks. The implication is that whatever switching processes were deployed in advance of the switch trials must have been fully effective, in that no further improvement in performance was observed on repeat trials, in terms of either accuracy or speed. In fact, in the case of the visual task there was a slight slowing of performance on repeat trials that suggested that anticipatory resources were not as effectively deployed as they had been on the preceding switch trials. This latter finding is consistent with the recorded physiology in that there was clearly less alpha desynchronisation on visual-repeat trials than on visual-switch trials, suggesting less effective engagement of visual cortical regions.

5A with 4I and K), and simply consisted in a time-independent reduction of the current. In order to test the physiological relevance of our voltage-clamp results, experiments were also performed in current clamp in the presence of the synaptic blockers mentioned above, but in the absence of TEA. Action potentials were evoked by a short (3-ms) depolarizing pulse (50–400 pA). Under these conditions, only ω-conotoxin (1 μm) was able to reduce the amplitude of the BMI-sensitive mAHP (by 79.7 ± 15.7%; n = 6). Mibefradil (30 μm) was devoid of any effect (n = 5). When

we co-applied the two blockers, Selleckchem Lumacaftor the reduction in the amplitude of the mAHP amounted to 79.5 ± 14.4% (Fig. 6A; n = 5). The three experimental conditions (mibefradil alone, ω-conotoxin alone and co-application of the two agents) induced a differential block of the mAHP ( = 7.47, P = 0.0077, Kruskal–Wallis test). Both ω-conotoxin alone and co-application of ω-conotoxin and mibefradil produced find more a significantly larger effect than mibefradil alone (U = 2.74, P = 0.006 and U = 2.6, P = 0.009, respectively; Mann–Whitney test).

We next performed intracellular recordings in the current-clamp mode in DR neurons from adult rats to test the sensitivity of the mAHP to blockers. Concentration–inhibition curves were first constructed with apamin (n = 5). The IC50 of the peptide was 2.5 ± 0.7 nm (not shown) with a mean Hill Protirelin coefficient of 2.5. In addition, we tested the sensitivity of the mAHP to tamapin, whose IC50 was found to be 9 and 17 nm (n = 2; mean Hill coefficient was 3.6; not shown). Taken together, these results suggest (but do not prove) that SK3 subunits are the main components of the SK channels underlying the mAHP of DR neurons. We then performed the same pharmacological experiments as above using Ca2+-channel blockers. Superfusion of ω-conotoxin (1 μm) also markedly reduced the amplitude of the mAHP in adult DRN serotonergic neurons (n = 6; mean inhibition 83 ± 3%). Its effect developed progressively to reach a stable maximum after 8 min. In contrast, no modification of the mAHP was observed with either mibefradil (30 μm; n = 4) or nifedipine (20 μm;

n = 4; Fig. 6B and C). In addition, the effect of ω-conotoxin was again not increased by the co-application of mibefradil (n = 4; not shown). A mixed anova showed a highly significant interaction between time and groups (F = 5.46, P < 0.001). The effect of ω-conotoxin was significantly higher than that of the two other blockers (P < 0.001 in both cases). The previous results show that N-type channels are the major source of Ca2+ that activates SK channels underlying the mAHP. However, these results were obtained in neurons which were silent (i.e. action potentials were induced by depolarizing current injection). In vivo, 5-HT neurons are known to have a slow pacemaker-like firing, at least in anaesthetized animals (Jacobs & Fornal, 1991).