A 2 bp deletion in the mitochondrial ATP 6 gene responsible for the NARP (neuropathy, ataxia, and retinitis pigmentosa) syndrome.

Mitochondrial (mt) DNA-associated NARP (neurogenic muscle weakness, ataxia, and retinitis pigmentosa) syndrome is due to mutation in the MT-ATP6 gene. We report the case of a 18-year-old man who presented with deafness, a myoclonic epilepsy, muscle weakness since the age of 10 and further developed a retinitis pigmentosa and ataxia. The whole mtDNA analysis by next-generation sequencing revealed the presence of the 2 bp microdeletion m.9127-9128 del AT in the ATP6 gene at 82% heteroplasmy in muscle and to a lower load in blood (10-20%) and fibroblasts (50%). Using the patient's fibroblasts, we demonstrated a 60% reduction of the oligomycin-sensitive ATPase hydrolytic activity, a 40% decrease in the ATP synthesis and determination of the mitochondrial membrane potential using the fluorescent probe tetramethylrhodamine, ethyl ester indicated a significant reduction in oligomycin sensitivity. In conclusion, we demonstrated that this novel AT deletion in the ATP6 gene is pathogenic and responsible for the NARP syndrome.

Vemurafenib skin phototoxicity is indirectly linked to ultraviolet A minimal erythema dose decrease.

BACKGROUND: Vemurafenib, an anti-rapidly accelerated fibrosarcoma kinase B (BRAF) molecule, improves survival among patients with metastatic BRAF-mutated melanoma. Photosensitivity, a frequent cutaneous adverse effect induced by vemurafenib, can lead to cessation of treatment. OBJECTIVES: To investigate photosensitivity mechanisms in patients treated with vemurafenib for metastatic melanoma. METHODS: In a prospective study of 12 patients, photobiological explorations with measurements of ultraviolet A (UVA) minimal erythema dose (MED) and polychromatic MED were performed over 3 days in all 12 patients. UVA MED and polychromatic MED were also assessed for four patients before treatment. We then performed spectrophotometric analyses of (i) serum and faeces in these four patients, before and after introduction of vemurafenib; (ii) the lyophilized form of vemurafenib without excipient added; and (iii) the lyophilized form of vemurafenib added to serum and faeces before treatment. RESULTS: Photosensitivity was present in 92% of the patients. UVA MED was normal before treatment and decreased after treatment, while polychromatic MED remained normal. The same three peaks (210, 260 and 310 nm) were identified in the spectrum for UVB and UVC but not for UVA on spectrophotometric analyses for each condition (lyophilized vemurafenib; serum and faeces after introduction of vemurafenib; and lyophilized vemurafenib added to serum and faeces before treatment). The peaks were different before treatment. CONCLUSIONS: Our study confirms that photosensitivity under vemurafenib treatment was a UVA phototoxicity reaction, and our results suggest that a metabolite of vemurafenib rather than the parent molecule is involved in this phototoxicity.

Effect of lipopolysaccharide, cytokines, and catecholamines on brain natriuretic peptide release from human myocardium.

BACKGROUND: During sepsis and septic shock, elevated plasma concentrations of brain natriuretic peptide (BNP) have been reported but may be related to several underlying mechanisms. The aim of the present experimental study was to investigate the effect of lipopolysaccharide (LPS), tumor necrosis factor α (TNF-α), interleukin 1ß (IL-1ß), interleukin 6 (IL-6), dobutamine (Dobu), epinephrine (Epi), and norepinephrine (Nor) on BNP synthesis by atrial human myocardium in vitro. METHODS: After the approval of local ethics committee, right atrial appendages were obtained during cannulation for cardiac surgery and pinned in a isolated organ bath containing 15 ml of Tyrode's modified solution. Preparations were oxygenated, maintained at 36 ± 0.5°C and stimulated at a frequency of 1 Hz. A 60-min equilibration period was followed by 180-min exposure to 1 µM endothelin 1 (ET-1; n = 9), 20,000 pg/ml TNF-α (n = 10), 1000 pg/ml IL-1ß (n = 10), 5000 pg/ml IL-6 (n = 10), 10,000 pg/ml LPS (n = 10), 100 µM Epi (n = 9), 100 µM Nor (n = 10), and 100 µM Dobu (n = 8). No product was added in Control group (n = 10). Two BNP dosages were performed: the first after 60 min of stabilization and the second after 180 min of stimulation. Absolute and relative changes in BNP concentration were compared between groups. RESULTS: Exposure to ET-1 significantly increased BNP release as compared with Control group. Dobu, Epi, Nor, and LPS significantly increased BNP concentration but not TNF-α, IL-1ß, or IL-6. CONCLUSIONS: In vitro, LPS, Dobu, Epi, and Nor induced BNP synthesis by human atrial myocardium.

Prior suggestive symptoms in one-third of patients consulting for a "first" demyelinating event.

OBJECTIVE: To evaluate the prevalence of prior inflammatory events in patients consulting for a first inflammatory neurological event and improve early diagnosis of multiple sclerosis. METHODS: During the initial visit, the neurologist gave patients a self-administered questionnaire containing 72 questions regarding previous symptoms lasting >24 h. During the follow-up visit, the neurologist validated the symptoms and collected information about the current attack. RESULTS: The cohort included 178 patients (74% women, mean age (SD) 33.7 (10.1) years). The main reason for the initial visit was visual disturbance and sensory troubles in limbs. Mean (SD) global Expanded Disability Status Scale score was 1.4 (1.1), 46% of brains MRIs were positive according to Barkhof-Tintoré criteria, 41% had abnormal white blood cell count in cerebrospinal fluid and 71% had immunoglobin G oligoclonal bands. Prior symptoms suggestive of demyelination were reported by 79 patients (44%), validated by the neurologist for 70% (55 patients) and identified only by the neurologist in four patients. Sequelae were observed in 14 patients with validated prior symptoms (26%). The self-administered questionnaire showed an overall sensitivity of 93% and specificity of 80% for identifying patients with prior symptoms suggestive of demyelination. CONCLUSION: A patient-administered questionnaire subsequently validated by the neurologist demonstrated that 33% of patients consulting for a first demyelinating event had prior symptoms suggestive of central nervous system demyelination that had gone unnoticed, and almost 70% had either sequelae of prior demyelination or McDonald criteria for dissemination in space. Such a questionnaire could be a useful tool for earlier diagnosis of multiple sclerosis.

[Mitochondrial diseases in adults: An update]. / Les maladies mitochondriales de l'adulte : mise au point.

Mitochondrial diseases, characterized by a respiratory chain deficiency, are considered as rare genetic diseases but are the most frequent among inherited metabolic disorders. The complexity of their diagnosis is due to the dual control by the mitochondrial (mtDNA) and the nuclear DNA (nDNA), and to the heterogeneous clinical presentations; illegitimate association of symptoms should prompt the clinician to evoke a mitochondrial disorder. The goals of this review are to provide clinicians a better understanding of mitochondrial diseases in adults. After a brief overview on the mitochondrial origin and functions, especially their role in the energy metabolism, we will describe the genetic bases for mitochondrial diseases, then we will describe the various clinical presentations with the different affected tissues as well as the main symptoms encountered. Even if the new sequencing approaches have profoundly changed the diagnostic process, the brain imaging, the biological, the biochemical, and the histological explorations are still important highlighting the need for a multidisciplinary approach. While for most of the patients with a mitochondrial disease, only supportive and symptomatic therapies are available, recent advances in the understanding of the pathophysiological mechanisms have been made and new therapies are being developed and are evaluated in human clinical trials.

Interactions between ethylenediaminetetraacetic acid (EDTA) and iron absorption pathways, in the Caco-2 model.

UNLABELLED: EDTA is a well known enhancer of iron absorption; however, the precise way of absorption of iron ingested in presence of EDTA is not known; some data suggest it could use a passive, non regulated paracellular way. Iron (sulphate or gluconate) absorption by Caco-2 cells was assessed in presence of EDTA. EDTA did not change the apical uptake of iron; transport in the basal chamber increased by 98% for FeSO4 and 95% for Fe gluconate. By contrast, intracellular storage decreased by 31% for FeSO4 and 64% for Fe gluconate. In addition EDTA induced a significant increase of permeability of the cell monolayer assessed by a decrease of transepithelial electrical resistance: 314+/-34 Omegacm(-2) to 235+/-57 Omegacm(-2) for sulphate, 414+/-33 Omegacm(-2) to 223+/-36 Omegacm(-2) for gluconate; iron free control: 410+/-10 Omegacm(-2). CONCLUSIONS: These results suggest that in presence of EDTA iron absorption occurs mainly by the paracellular instead of the regulated cellular way, that could potentially enhance its toxicity.

In vitro and in vivo pharmacological profile of UFP-512, a novel selective delta-opioid receptor agonist; correlations between desensitization and tolerance.

BACKGROUND AND PURPOSE: Delta-opioid receptors (DOP receptors) could represent a novel target in the treatment of depressive disorders. To explore this new field of interest, the development of highly selective DOP receptor agonists is essential. UFP-512 [H-Dmt-Tic-NH-CH(CH2-COOH)-Bid], was recently shown to behave in vitro as a selective and potent DOP receptor agonist and to promote antidepressant- and anxiolytic-like effects in vivo (Vergura et al., 2007). Here, we have characterized the pharmacological properties of UFP-512 and established a link between desensitization and tolerance. EXPERIMENTAL APPROACH: Studies were performed in the human neuroblastoma SK-N-BE cells to establish i) binding parameters for UFP-512 ii) signalling pathways activated after acute and chronic treatment iii) regulation (phosphorylation and trafficking) of human DOP (hDOP) receptors after sustained activation by UFP-512. In vivo, we studied UFP-512-induced antidepressant-like effects after acute or chronic treatment in the mouse forced swimming test. KEY RESULTS: In vitro, UFP-512 was a high affinity agonist for DOP receptors. While UFP-512 induced marked phosphorylation of DOP receptors on Ser363, we observed a low desensitization of the cAMP pathway, associated with receptor endocytosis and recycling without any reduction on extracellular signal-regulated protein kinase 1/2 activation. In vivo, acute administration of UFP-512 produced an antidepressant-like effect, without any sign of tolerance after chronic administration. CONCLUSIONS AND IMPLICATIONS: There was a correlation between weak desensitization, significant internalization and recycling of the human DOP receptors and lack of tolerance to UFP-512. This suggests that this compound would be a promising drug prototype for exploring innovative treatments for mood disorders.

Evaluation of cardiac troponin I stability in blood sample using the AccuTnI+3 assay.

OBJECTIVE: Troponins are considered as the biomarkers of choice to highlight cardiac injury in emergency departments, but are also valuable to detect cardiac injury in a non-emergency setting. In this latter case, transport of blood samples to laboratories often exceeds the manufacturer's recommendations (<2 h between vein puncture and analysis for the Beckman Coulter AccuTnI+3 assay). We aim to evaluate in vitro the stability of troponin Ic (cTnI) at two intervals (<2 h and at 4 h) over a wide range of concentrations using the Beckman Coulter AccuTnI+3 assay. PATIENTS AND METHODS: For each of the 95 patients included in this study, we analyzed the first blood sample with a time of transport <2 h, and the second sample after 4 h from vein puncture. We then calculated the correlation between the two periods of analysis and evaluated the bias by a Bland-Altman test. RESULTS: Taking into account of our analytical reproducibility, we did not observe any significant differences in cTnI values between <2 h and 4 h. CONCLUSIONS: The time between vein puncture and analysis of cTnI can be extended to 4 h.

Effects of glycaemic variability on cardiac remodelling after reperfused myocardial infarction: Evaluation of streptozotocin-induced diabetic Wistar rats using cardiac magnetic resonance imaging.

AIMS: In addition to hyperglycaemia, glycaemic variability seems to be associated with poor outcomes after acute myocardial infarction. This study explored the impact of glycaemic variability in diabetic Wistar rats subjected to myocardial ischaemia/reperfusion. METHODS: Animals with streptozotocin-induced diabetes received insulin either to maintain stable hyperglycaemia (Dh group) or to generate glycaemic variability (Dv). After experimental myocardial ischaemia/reperfusion was surgically induced, 7T cardiac magnetic resonance imaging (CMR) was performed at weeks 1 (w1) and 3 (w3). RESULTS: Twenty-six rats were randomized [sham group (S): n=5; control group (C): n=7; Dh group: n=6; and Dv group: n=8]. The mean amplitude of glucose reflecting glycaemic variability was higher in the Dv than in the Dh group (9.1±2.7mmol/L vs 5.9±1.9mmol/L; P<0.05). CMR assessment at w3 revealed ventricular enlargement in both Dh and Dv groups compared with the C and S groups (end-diastolic volume: 1.60±0.22 and 1.36±0.30mL/kg compared with 1.11±0.13 and 0.87±0.11mL/kg, respectively; P<0.05). Circumferential strain was altered between w1 and w3 in the remote area only in the Dv group, resulting in a lower value in this group than in the S, C and Dh groups (-0.11±0.01 vs -0.17±0.05, -0.15±0.03 and -0.16±0.03, respectively; P<0.05). In addition, at w3, oedema was also higher in the remote area in the Dv than in the C group (18.3±4.9ms vs 14.5±1.7ms, respectively; P<0.05). CONCLUSION: In the context of experimental myocardial ischaemia/reperfusion, our results suggest that glycaemic variability might have a potentially deleterious impact on myocardial outcomes beyond the classical glucose metrics.

Pharmacological delta1- and delta2-opioid receptor subtypes in the human neuroblastoma cell line SK-N-BE: no evidence for distinct molecular entities.

The two pharmacological delta-opioid receptor subtypes, delta1 and delta2, have been defined on the basis of pharmacological tools but remain to be characterized at the molecular level, since only a single cDNA has been cloned. The present study aimed to investigate the pharmacological properties of delta1- and delta2-opioid subtypes expressed in the human neuroblastoma cell line SK-N-BE and to characterize their putative corresponding mRNAs. Binding experiments using "selective" delta1- and delta2-opioid agonists and antagonists revealed the presence of two binding sites, demonstrating the presence of these delta1-opioid subtypes as they were previously described. The activation of these pharmacological subtypes by the selective agonists induced the incorporation of [alpha-(32)P]azidoanilide-GTP into Galpha(i2)/Galpha(0) subunits with the same efficiency and potency and inhibited adenosine 3', 5'-cyclic monophosphate (cAMP) accumulation with similar efficiency, while their sustained activation for 15 min induced a cross-desensitization. The "selective" delta1 and delta2 antagonists, 7-benzylidenenaltrexone and naltrindole benzofuran, respectively, were found to be as potent in blocking the inhibition of cAMP accumulation induced by both [D-Pen(2,5)]enkephalin and Tyr-D-Ala-Phe-Asp-Val-Val-Gly-NH(2). The possibility that delta-opioid subtypes could arise from alternative splicing was ruled out by reverse transcription-polymerase chain reaction (RT-PCR) experiments and the sequencing of PCR products, which revealed the presence of a single transcript encoding for the delta-opioid receptor. Different possibilities which could account for the delta-opioid receptor heterogeneity observed in the SN-N-BE cell line are discussed.

Differential desensitization of human delta-opioid receptors by peptide and alkaloid agonists.

The efficacy of different opioid agonists to induce acute desensitization of the human delta-opioid receptor-mediated inhibition of cAMP accumulation was investigated in the neuroblastoma cell line SK-N-BE, which endogenously expresses these receptors. While etorphine, a non-selective alkaloid agonist, caused 50% desensitization after a 30-min incubation, the same treatment in the presence of the selective peptide agonists, DPDPE ([D-Pen2,D-Pen5]enkephalin) and deltorphin I (Tyr-D-Ala-Phe-Asp-Val-Val-Gly), almost totally desensitized the delta-opioid receptor-mediated inhibition of adenylyl cyclase. When SK-N-BE cells were prechallenged either with alkaloid or peptide agonist, we observed a cross-desensitization that was less marked when cells were pretreated with peptide agonists and then challenged with etorphine. Taken together, these results demonstrate that human delta-opioid receptors are differentially desensitized by alkaloid and peptide agonists.

[Profile and evolution of irritable bowel syndrome. Prospective national epidemiological study of 1301 patients followed-up for 9 months in Gastroenterology. Groupe d'Etude Nationale sur le Syndrome de l'Intestin Irritable (SII)]. / Profil et évolution du syndrome de l'intestin irritable. Etude épidémiologique nationale prospective chez 1,301 consultants en Gastroentérologie suivis 9 mois. Groupe d'Etude Nationale sur le Syndrome de l'Intestin Irritable (SII).

From March to October 1989, 237 French gastroenterologists included 1,301 patients referred for irritable bowel syndrome in a 9-month epidemiological survey based on questionnaires and monthly auto-evaluation. In the patient population, the high preponderance of women (sex ratio: 2.33), the high prevalence of cholecystectomy (9%), appendectomy (53%) and an association with at least one symptom of non-ulcer dyspepsia (70%) were observed. Fifty per cent of the patients completed the 9-month follow-up period; among them, 60% declared an improvement in their symptoms, but only 30% in their quality of life and independently of the clinical course. This study suggests that symptoms and quality of life in patients consulting for irritable bowel syndrome should be taken into account separately, both in daily practice and in therapeutic evaluation.

[Descriptive study of digestive functional symptoms in the French general population]. / Etude descriptive des symptômes fonctionnels digestifs dans la population générale française.

UNLABELLED: To study the prevalence of "reported" functional digestive symptoms (FDS) in terms easily understood by the general population without resorting to predefined concepts of functional syndromes, and to assess FDS impact on public health, a sample survey has been carried out between September and December 1995. METHODS: Four thousand eight hundred and seventeen subjects representative of the French general population aged 15 years or more filled in a questionnaire describing their digestive disorders. RESULTS: Seventy percent of the subjects had digestive complaints, 9% being related to a presumably organic disease, and 61% attributed to FDS. Twenty-seven % of the subjects claimed to be inconvenienced by their FDS, whereas 34% seemed not to feel any inconvenience. Among FDS, gas emission was the most frequent symptom (59%), followed by stomach ache and/or digestive pain (48%), flatulence (47%), bad digestion sensations (40%), constipation (35%), aerophagia (29%), bad breath (22%), incomplete evacuation of stools (19%). FDS had lasted from 6 months to 5 years in 38%, and over 5 years in 52%. In the subgroup of subjects inconvenienced by FDS (27%), 9% consulted and 18% did not, whereas in the subgroup not inconvenienced, 3% consulted and 31% did not. Altogether, 26% of the subjects followed a prescription or self medication treatment; 35% were not treated. Some explanatory variables appeared to be associated with the onset of inconvenience: the associations pain and bad digestion, flatulence and aerophagia, incomplete evacuation and nervous or presumably organic origin of FDS, age, stress, FDS frequency. Duration of symptoms, age above 65 years, digestive pain, presumably organic origin, and FDS frequency were associated with the need to consult. This descriptive, pragmatic survey shows the widespread prevalence of FDS, affecting 28 million French people. Functional digestive disorders in the "academic" meaning constitute only a limited subset. FDS lead to major health care consumption. Their impact on public health is undoubtedly greater than the estimates derived from studies designed in accordance with conventional nosological categories.

[Familial idiopathic striato-pallido-dentate calcifications: clinical and brain imaging study in a family]. / Calcifications striato-pallido-dentelées idiopathiques familiales. Etude clinique et en imagerie d'une famille et revue de la littérature.

Familial idiopathic basal ganglia calcification (FIBGC) is a rare condition and its pathophysiology has not so far been elucidated. We report the results of a clinical study in two patients of a family affected with FIBGC. Brain imaging with 18-FDG-PET was performed in one. Psychiatric and cognitive troubles were the main clinical symptoms. Basal ganglia calcifications were associated with white matter lesions. The PET study performed in one patient revealed a striatal and a posterior cingulate hypometabolism. Posterior cingulate gyrus is involved in episodic memory processing, and could be involved in episodic memory deficit observed in this patient. These results suggest that a cortical dysfunction could be associated to the disease. The underlying mechanism, that could be a neuronal loss, a cortical deafferentation or an alteration of synaptic transmission, remains to be elucidated.

[Prevalence of uterine fibroids in France and impact on quality of life: results of a survey among 2500 women between 30-55 years]. / Prévalence du fibrome utérin en France et impact sur la qualité de vie à partir d'une enquête menée auprès de 2500 femmes de 30-55ans.

OBJECTIVES: To evaluate the prevalence of symptomatic uterine fibroid in France, related symptoms and its impact on quality of life. MATERIALS AND METHODS: Online prospective survey from December 2012 to February 2013 using two questionnaires on 2498 women aged between 30 and 55 years old and representatives of the feminine population. RESULTS: On the studied population, 220 women presented a symptomatic uterine fibroid representing a prevalence of 8.8%. 163 women (73.7%) presented bleedings associated or not with pain, and 58 (26.3%) suffered from pain only. For 77 (34.9%) women, the symptoms preceded the diagnostic and the mean delay between first symptoms and diagnostic was 2.22 years (ET : 3.56). At the time of the survey, 144 (65%) women with symptomatic uterine fibroid were followed by a physician (a gynecologist in 91.6% (n=132)), and 110 (49.7%) evaluated their pain as severe or extremely severe and 178 (80.6%) were bothered by their symptoms in their everyday life. CONCLUSION: This analysis shows a strong prevalence for symptomatic uterine fibroid with a major impact on the quality of life.

Remifentanil produces cross-desensitization and tolerance with morphine on the mu-opioid receptor.

Remifentanil is a powerful mu-opioid (MOP) receptor agonist used in anaesthesia with a very short half-life. However, per-operative perfusion of remifentanil was shown to increase morphine consumption during post-operative period to relieve pain. In the present study, we aimed to describe the cellular mechanisms responsible for this apparent reduction of morphine efficacy. For this purpose, we first examined the pharmacological properties of both remifentanil and morphine at the MOP receptor, endogenously expressed in the human neuroblastoma SH-SY5Y cell line, to regulate adenylyl cyclase and the MAP kinase ERK1/2 pathway, their potency to promote MOP receptor phosphorylation, arrestin 3-CFP (cyan fluorescent protein) recruitment and receptor trafficking during acute and sustained exposure. In the second part of this work, we studied the effects of a prior exposure of remifentanil on morphine-induced inhibition of cAMP accumulation, activation of ERK1/2 and analgesia. We showed that sustained exposure to remifentanil promoted a rapid desensitization of opioid receptors on both signalling pathways and a pretreatment with this agonist reduced signal transduction produced by a second challenge with morphine. While both opioid agonists promoted Ser(375) phosphorylation on MOP receptor, remifentanil induced a rapid internalization of opioid receptors compared to morphine but without detectable arrestin 3-CFP translocation to the plasma membrane in our experimental conditions. Lastly, a cross-tolerance between remifentanil and morphine was observed in mice using the hot plate test. Our in vitro and in vivo data thus demonstrated that remifentanil produced a rapid desensitization and internalization of the MOP receptor that would reduce the anti-nociceptive effects of morphine.

Development and implementation of standardized respiratory chain spectrophotometric assays for clinical diagnosis.

Diversity of respiratory chain spectrophotometric assays may lead to difficult comparison of results between centers. The French network of mitochondrial diseases diagnostic centers undertook comparison of the results obtained with different protocols in the French diagnostic centers. The diversity of protocols was shown to have striking consequences, which prompted the network to undertake standardization and optimization of the protocols with respect to clinical diagnosis, i.e. high velocity while maintaining linear kinetics relative to time and enzyme concentration. Assays were set up on animal tissues and verified on control human muscle and fibroblasts. Influence of homogenization buffer and narrow range of optimal concentration of phosphate, substrate and tissue were shown. Experimental data and proposed protocols have been posted on a free access website. Their subsequent use in several diagnostic centers has improved consistency for all assays.

A novel mutation 3090 G>A of the mitochondrial 16S ribosomal RNA associated with myopathy.

We describe a young woman who presented with a progressive myopathy since the age of 9. Spectrophotometric analysis of the respiratory chain in muscle tissue revealed combined and profound complex I, III, II+III, and IV deficiency ranging from 60% to 95% associated with morphological and histochemical abnormalities of the muscle. An exhaustive screening of mitochondrial transfer and ribosomal RNAs showed a novel G>A substitution at nucleotide position 3090 which was detected only in urine sediment and muscle of the patient and was not found in her mother's blood cells and urine sample. We suggest that this novel de novo mutation in the 16S ribosomal RNA, a nucleotide which is highly conserved in different species, would impair mitochondrial protein synthesis and would cause a severe myopathy.

The delta-opioid receptor regulates activity of ryanodine receptors in the human neuroblastoma cell line SK-N-BE.

Recent studies have demonstrated that opioid agonists affect the cytosolic Ca2+ concentration ([Ca2+]i) either by regulating plasma membrane Ca(2+)-channel activity or by mobilizing intracellular Ca2+ stores. The present report documents the [Ca2+]i increase induced by opioid agonists in a human neuroblastoma cell line, SK-N-BE, expressing delta-opioid receptors. In the presence, as well as in the absence, of extracellular Ca2+, opioid agonists enhanced significantly [Ca2+]i, whereas carbachol, known to mobilize specifically inositol 1,4,5-trisphosphate-sensitive intracellular Ca2+ stores, acted only in the presence of extracellular Ca2+. The opioid-induced increase in [Ca2+]i was not affected by treatments modifying the trimeric Gl, Go, and Gs protein transduction mechanisms or the activity of adenylyl cyclase. The Ca(2+)-ATPase pump-inhibiting sesquiterpene lactone, thapsigargin, did not modify the opioid-induced [Ca2+]i response, whereas it abolished the effects of carbachol. The Ryana speciosa alkaloid, ryanodine, at concentrations known to block endoplasmic reticulum ryanodine receptors, decreased significantly the response to opioids without affecting the effects of carbachol. Thus, our results suggest that, in SK-N-BE cells, delta-opioid receptors mobilize Ca2+ from intracellular ryanodine-sensitive stores and the mechanism involved is independent of Gl/Go Gs proteins and protein kinase A activation.