Radiofrequency ablation versus surgical resection for multiple HCCs meeting the Milan criteria: propensity score analyses of 10-year therapeutic outcomes.

AIM: To assess the long-term therapeutic outcomes of radiofrequency ablation (RFA) versus surgical resection (SR) as a first-line treatment for patients meeting the Milan criteria with multiple hepatocellular carcinomas (HCCs). MATERIALS AND METHODS: This retrospective study was approved by the institutional review board and the requirement for informed consent was waived. Between January 2004 and December 2009, among 3,441 patients with treatment-naive HCCs, 88 patients meeting the Milan criteria with multiple HCCs (Barcelona Clinic Liver Cancer [BCLC] A stage) who underwent either RFA (n=62) or SR (n=26) were included. Recurrence-free survival (RFS) and overall survival (OS) rates were compared by using propensity score matching. In addition, multivariate analysis was performed for assess the prognostic factor. RESULTS: Matching yielded 20 matched pairs of patients. In the two matched groups, the RFS rates were 30% and 30% at 5- and 10-years, respectively, in the RFA group and 60% and 48.6% in the SR group (p=0.054). The corresponding OS rates were 63.3% and 46.1% in the RFA group and 100% and 73.6% in the SR group, respectively (p=0.061). In multivariate analysis, treatment type was independently associated with RFS (hazard ratio [HR]=0.51; p=0.043) whereas it was not a statistically significant factor for OS (HR=0.50; p=0.088). CONCLUSION: In patients meeting the Milan criteria with multiple HCCs (BCLC A stage), SR may provide better RFS compared to RFA.

Temporal variability of glucocorticoid receptor activity is functionally important for the therapeutic action of fluoxetine in the hippocampus.

Previous studies have shown inconsistent results regarding the actions of antidepressants on glucocorticoid receptor (GR) signalling. To resolve these inconsistencies, we used a lentiviral-based reporter system to directly monitor rat hippocampal GR activity during stress adaptation. Temporal GR activation was induced significantly by acute stress, as demonstrated by an increase in the intra-individual variability of the acute stress group compared with the variability of the non-stress group. However, the increased intra-individual variability was dampened by exposure to chronic stress, which was partly restored by fluoxetine treatment without affecting glucocorticoid secretion. Immobility in the forced-swim test was negatively correlated with the intra-individual variability, but was not correlated with the quantitative GR activity during fluoxetine therapy; this highlights the temporal variability in the neurobiological links between GR signalling and the therapeutic action of fluoxetine. Furthermore, we demonstrated sequential phosphorylation between GR (S224) and (S232) following fluoxetine treatment, showing a molecular basis for hormone-independent nuclear translocation and transcriptional enhancement. Collectively, these results suggest a neurobiological mechanism by which fluoxetine treatment confers resilience to the chronic stress-mediated attenuation of hypothalamic-pituitary-adrenal axis activity.

Radiofrequency ablation and transarterial chemoembolisation as first-line treatment for recurrent hepatocellular carcinoma or isolated intrahepatic recurrent hepatocellular carcinoma in transplanted livers.

AIM: To evaluate the efficacy of radiofrequency ablation (RFA) and transarterial chemoembolisation (TACE) as a first-line treatment for isolated intrahepatic recurrent hepatocellular carcinoma (IIR-HCC) after liver transplantation (LT). MATERIALS AND METHODS: This retrospective study was approved by the institutional review board. Between January 2005 and January 2015, 588 consecutive patients underwent LT for the treatment of HCC. Among them, 27 patients with IIR-HCCs after LT who were treated with RFA (n=6) or TACE (n=21) as a first-line treatment were retrospectively included in this study. Disease-free and overall survival rates were estimated using the Kaplan-Meier method. Risk factors affecting these outcomes were assessed with Cox regression models. RESULTS: Except for the total number of recurrent tumours and time-to-tumour recurrence after LT, baseline characteristics were not significantly different between the groups. The 2-year disease-free survival rates for RFA and TACE (20% versus 14%, respectively; p=0.180) and 4-year overall survival rates (33% versus 25%, respectively; p=0.065) were not significantly different between groups. In addition, the types of treatment were not associated with disease-free or overall survival in multivariate analyses. CONCLUSION: TACE may be an effective treatment comparable to RFA in patients with IIR-HCC after LT when RFA is not feasible.

Percutaneous ultrasonography-guided radiofrequency ablation of hepatocellular carcinomas: usefulness of image fusion with three-dimensional ultrasonography.

AIM: To evaluate the usefulness of fusion imaging with real-time ultrasonography (US) and three-dimensional (3D) US for the guidance of radiofrequency ablation (RFA) of hepatocellular carcinomas (HCCs) 2-5 cm in diameter. MATERIALS AND METHODS: This study was conducted as a retrospective cohort study. It was approved by the institutional review board and informed consent was waived. During percutaneous RFA of HCCs, targeting was performed under conventional fusion imaging guidance, whereas monitoring and controlling were conducted under fusion with 3D US guidance. Technical success, technique effectiveness, incidence of major complications, and local tumour progression rate were evaluated. According to tumour size (small: <3 cm versus medium: 3-5 cm), the roundness indexes of the ablation zones and local tumour progression rates were compared. RESULTS: There were 29 small-sized HCCs (2.5 ± 0.3 cm) and 17 medium-sized HCCs (3.4 ± 0.5 cm). All RFA procedures were performed in a single RFA session. Both the technical success and technique effectiveness rates were 100%. One patient with medium-sized HCC developed a hepatic abscess (n = 1) as a major complication. The local tumour progression rate was 8.7% (4/46) with a mean follow-up period of 18.2 months. The roundness indexes of the ablation zone were not significantly different between small- and medium-sized HCCs, and the local tumour progression rates were also not significantly different between the two groups [3.4% (1/29) versus 17.6% (3/17); p = 0.135]. CONCLUSION: Image fusion with real-time US and 3D US is useful for the guidance of percutaneous RFA for HCCs 2-5 cm in diameter.

Local tumour progression after loco-regional therapy of hepatocellular carcinomas: value of fusion imaging-guided radiofrequency ablation.

AIM: To assess the value of fusion imaging of real-time ultrasonography (US) with liver computed tomography (CT)/magnetic resonance imaging (MRI) images for planning US of radiofrequency ablation (RFA) in improving conspicuity of the lesions and reducing false-positive detection of local tumour progression (LTP) found after transcatheter arterial chemoembolization (TACE) or RFA of hepatocellular carcinoma (HCC). MATERIALS AND METHODS: This study was approved by the institutional review board and informed consent was waived. Fifty patients with LTP (mean ± SD, 1.5 ± 0.6 cm; range 0.5-3 cm) detected at follow-up CT or MRI were included. Planning US was performed by two radiologists using conventional US first and fusion imaging later in the same session. False-positive detection rates were assessed using conventional US based on the results of fusion imaging. The number cases of initially invisible tumours on conventional US that became visible after image fusion were also evaluated. The true-positive detection rate and conspicuity scores of the index tumours were compared between conventional US and fusion imaging. RESULTS: On conventional US, 40 (80%) out of 50 HCCs with LTP were identified. However, the false-positive detection rate of conventional US was 12.5% (5/40). Out of 10 initially invisible HCCs with LTP on conventional US, six (60%) became visible after image fusion. The true-positive detection rate on conventional US was 70% (35/50), whereas it was increased to 92% (46/50) after image fusion (p = 0.0026). CONCLUSION: Fusion imaging can improve the conspicuity of lesions and reduce the false-positive detection of LTP after TACE or RFA.

Percutaneous radiofrequency ablation of hepatic tumours: factors affecting technical failure of artificial ascites formation using an angiosheath.

AIM: To evaluate the technical feasibility of artificial ascites formation using an angiosheath before percutaneous radiofrequency ablation (RFA) for hepatic tumours and to determine predictive factors affecting the technical failure of artificial ascites formation. MATERIALS AND METHODS: This retrospective study was approved by the institutional review board. One hundred and thirteen patients underwent percutaneous RFA of hepatic tumours after trying to make artificial ascites using an angiosheath to avoid collateral thermal damage. The technical success rate of making artificial ascites using an angiosheath and conversion rate to other techniques after initial failure of making artificial ascites were evaluated. The technical success rate for RFA was assessed. In addition, potential factors associated with technical failure including previous history of transcatheter arterial chemoembolization (TACE) or RFA, type of abdominal surgery, and adjacent perihepatic structures were reviewed. Predictive factors for the technical failure of artificial ascites formation were analysed using multivariate analysis. RESULTS: The technical success rates of artificial ascites formation by angiosheath and that of RFA were 84.1% (95/113) and 97.3% (110/113), respectively. The conversion rate to other techniques after the failure of artificial ascites formation using an angiosheath was 15.9% (18/113). Previous hepatic resection was the sole independent predictive factor affecting the technical failure of artificial ascites formation (p<0.001, odds ratio = 29.03, 95% confidence interval: 4.56-184.69). CONCLUSION: Making artificial ascites for RFA of hepatic tumours using an angiosheath was technically feasible in most cases. However, history of hepatic resection was a significant predictive factor affecting the technical failure of artificial ascites formation.

First-line radiofrequency ablation with or without artificial ascites for hepatocellular carcinomas in a subcapsular location: local control rate and risk of peritoneal seeding at long-term follow-up.

AIM: To compare the long-term local control of hepatocellular carcinoma (HCC) and risk of peritoneal seeding via percutaneous radiofrequency ablation (RFA) using artificial ascites with those of RFA without artificial ascites. MATERIALS AND METHODS: The Institutional Review Board approved this retrospective study. From April 2005 to February 2008, 160 patients (121 men, 39 women; age range 36-79 years) with a single subcapsular HCC (mean size 2.19 cm) were treated with ultrasonography-guided percutaneous RFA as a first-line therapy. Forty-four patients were treated with RFA using artificial ascites, whereas the other 116 patients were treated without artificial ascites. The cumulative local tumour progression (LTP) and peritoneal seeding were compared in both groups using follow-up computed tomography (CT). Cumulative LTP rates were analysed using the Kaplan-Meier method and the log-rank test. Risk of peritoneal seeding was investigated by means of multivariate analysis. RESULTS: The overall median follow-up period was 52.5 months (range 13-76 months). The 1, 2, 4, and 6 year cumulative LTP rates were 17.1, 27.6, 35.2, and 35.2%, respectively, in the group with artificial ascites, and 8, 15.2, 26.6, and 34.4% in the group without artificial ascites, without significant difference (p = 0.332). The rates of peritoneal seeding were 6.8% (3/44) in the group with artificial ascites and 2.6% (3/116) in the group without artificial ascites, a non-significant difference (p = 0.347). The biopsy prior to RFA was the independent risk factor of peritoneal seeding regardless of the use of artificial ascites. CONCLUSION: Long-term local tumour control and risk of peritoneal seeding were comparable for RFA with or without artificial ascites when used as a first-line therapy for subcapsular HCC.

Volumetric feedback ablation of uterine fibroids using magnetic resonance-guided high intensity focused ultrasound therapy.

OBJECTIVE: The purpose of this prospective multicenter study was to assess the safety and technical feasibility of volumetric Magnetic Resonance-guided High Intensity Focused Ultrasound (MR-HIFU) ablation for treatment of patients with symptomatic uterine fibroids. METHODS: Thirty-three patients with 36 fibroids were treated with volumetric MR-HIFU ablation. Treatment capability and technical feasibility were assessed by comparison of the Non-Perfused Volumes (NPVs) with MR thermal dose predicted treatment volumes. Safety was determined by evaluation of complications or adverse events and unintended lesions. Secondary endpoints were pain and discomfort scores, recovery time and length of hospital stay. RESULTS: The mean NPV calculated as a percentage of the total fibroid volume was 21.7%. Correlation between the predicted treatment volumes and NPVs was found to be very strong, with a correlation coefficient r of 0.87. All patients tolerated the treatment well and were treated on an outpatient basis. No serious adverse events were reported and recovery time to normal activities was 2.3 ± 1.8 days. CONCLUSION: This prospective multicenter study proved that volumetric MR-HIFU is safe and technically feasible for the treatment of symptomatic uterine fibroids. KEY POINTS: • Magnetic-resonance-guided high intensity focused ultrasound allows non-invasive treatment of uterine fibroids. • Volumetric feedback ablation is a novel technology that allows larger treatment volumes • MR-guided ultrasound ablation of uterine fibroids appears safe using volumetric feedback.

Differentiating mass-forming intrahepatic cholangiocarcinoma from atypical hepatocellular carcinoma using gadoxetic acid-enhanced MRI.

AIM: To examine the differential features of mass-forming intrahepatic cholangiocarcinoma (ICC) from atypical hypovascular hepatocellular carcinoma (HCC) on gadoxetic acid-enhanced magnetic resonance imaging (MRI). MATERIALS AND METHODS: The institutional review board approved this retrospective study and waived informed patient consent. Seventy patients with pathologically proven ICCs (35) and hypovascular atypical HCCs (35) who had undergone preoperative gadoxetic acid-enhanced MRI were enrolled in this study. Images were analysed for the shape of the lesions and presence of hyperintensity on the T1-weighted image (T1WI) and hypo- or hyperintense areas on the T2-weighted image (T2WI). In addition, images were analysed for the presence of linear hyperintensity or multifocal, tiny, hyperintense foci on T2WI and the presence of rim enhancement during early dynamic phases and a central enhancement with a hypointense rim (target appearance) on the 10 and 20 min hepatobiliary phase images. The significance of these findings was determined by the X(2) test. RESULTS: Univariate analysis revealed that the following significant parameters favour ICC or hypovascular HCC; the presence of T2 hypo- and hyperintense areas and target appearance on the 10 min hepatobiliary phase images favour ICC, and the presence of T2 linear hyperintensity and T2 multifocal hyperintense foci favour hypovascular HCC (p < 0.05). Multivariate analysis revealed that only target appearance on the 10 min hepatobiliary phase was predictive of ICC (p = 0.002) as 30 ICCs (85.7%) showed this feature. However, the target appearance was also observed in all six scirrhous HCCs. CONCLUSION: A target appearance on the 10 min hepatobiliary phase images is the best predictor for identifying mass-forming ICC at gadoxetic acid-enhanced MRI.

The JAK2 V617F mutation in de novo acute myelogenous leukemias.

A missense somatic mutation in JAK2 gene (JAK2 V617F) has recently been reported in chronic myeloproliferative disorders, including polycythemia vera, essential thrombocythemia and myelofibrosis with myeloid metaplasia, strongly suggesting its role in the pathogenesis of myeloid disorders. As activation of JAK2 signaling is occurred in other malignancies as well, we have analysed 558 tissues from common human cancers, including colon, breast and lung carcinomas, and 143 acute adulthood leukemias by polymerase chain reaction -- single strand conformation polymorphism analysis. We found three JAK2 mutations in the 113 acute myelogenous leukemias (AMLs) (2.7%), but none in other cancers. The mutations consisted of two V617F mutations and one K607N mutation. None of the AML patients with the JAK2 V617F mutation had a history of previous hematologic disorders. This is the first report on the JAK2 gene mutation in AML, and the data indicated that the JAK2 gene mutation may not only contribute to the development of chronic myeloid disorders, but also to some AMLs.

Impaired long-term memory and long-term potentiation in N-type Ca2+ channel-deficient mice.

Voltage-dependent N-type Ca(2+) channels, along with the P/Q-type, have a crucial role in controlling the release of neurotransmitters or neuromodulators at presynaptic terminals. However, their role in hippocampus-dependent learning and memory has never been examined. Here, we investigated hippocampus-dependent learning and memory and synaptic plasticity at hippocampal CA3-CA1 synapses in mice deficient for the alpha(1B) subunit of N-type Ca(2+) channels. The mutant mice exhibited impaired learning and memory in the Morris water maze and the social transmission of food preference tasks. In particular, long-term memory was impaired in the mutant mice. Interestingly, among activity-dependent long-lasting synaptic changes, theta burst- or 200-Hz-stimulation-induced long-term potentiation (LTP) was decreased in the mutant, compared with the wild-type mice. This type of LTP is known to require brain-derived neurotrophic factor (BDNF). It was found that both BDNF-induced potentiation of field excitatory postsynaptic potentials and facilitation of the frequency of miniature excitatory postsynaptic currents (mEPSCs) were reduced in the mutant. Taken together, these results demonstrate that N-type Ca(2+) channels are required for hippocampus-dependent learning and memory, and certain forms of LTP.

The role of c-Myc and heat shock protein 70 in human hepatocarcinoma Hep3B cells during apoptosis induced by prostaglandin A2/Delta12-prostaglandin J2.

Prostaglandin (PG) A2 (PGA2) and Delta12-PGJ2 have potent antiproliferative activity on various tumor cell growths in vitro. In this study, we investigated the mechanism of PGA2/Delta12-PGJ2-mediated apoptosis, including intracellular apoptosis-related genes in human hepatocarcinoma Hep3B cells. Hep3B cells treated with PGA2/Delta12-PGJ2 showed that a time-dependent DNA fragmentation characterized by marked apoptosis and the elevation of c-myc mRNA expression. In proportion to the increased c-myc gene transcription, heat shock protein 70 (hsp70) mRNA was induced from 1 to 24 h after PGA2/Delta12-PGJ2 treatment. The transfection of c-myc antisense oligomers in Hep3B cells significantly delayed the induction of HSP70 expression and blocked formation of DNA fragmentation by PGA2/Delta12-PGJ2. Moreover, overexpressed HSP70 showed an increased resistance to apoptosis by PGA2/Delta12-PGJ2 treatment. These results demonstrated that the decreased survival in response to PGA2/Delta12-PGJ2 was causally related to the amount of c-myc and the induction of c-myc regulated the elevation of HSP70 which have been known to correlate with a resistance to apoptosis.

Costochondritis. A prospective analysis in an emergency department setting.

BACKGROUND: Costochondritis (CC) is a common, but poorly understood condition among patients with chest wall pain. We have prospectively analyzed distinctive features of patients presenting to the emergency department with chest pain and CC. METHODS: Patients with a chief complaint of chest pain, not due to trauma, fever, or malignancy, were prospectively evaluated for the presence of CC and compared with another chest pain group without CC. RESULTS: Of 122 consecutive patients studied, 36 had CC (30%) and in 17 the pain induced reproduced the original one (15%). Women made up 69% of the patients with CC (vs 31% of control subjects) and Hispanics 47% (vs 24% of control subjects). Only three patients (8%) with CC met the American College of Rheumatology criteria for fibromyalgia, while none of the control subjects did. Widespread pain was more common in the CC group (42% vs 5%). The mean sedimentation rate in the CC group was 44 +/- 31 mm/h vs 41 +/- 31 mm/h in the control group. The acute myocardial infarction rate was 6% in the CC group vs 28% in the control group. Rheumatoid arthritis and osteoarthritis were diagnosed in three and two patients, respectively, of 32 patients with CC cases. One year later, 11 (55%) of 21 patients with CC were still suffering from chest pain, but only one third still had definite CC. CONCLUSIONS: Costochondritis is common among patients with chest pain in an emergency department setting, with a higher frequency among women and Hispanics. It is associated with fibromyalgia in only a minority of cases. Patients with CC appear to have a lower frequency of acute myocardial infarction. Spontaneous resolution is seen in most cases at 1 year.

Hepatic radiofrequency ablation: in vivo and ex vivo comparisons of 15-gauge (G) and 17-G internally cooled electrodes.

OBJECTIVE: To compare the performance of the 15-G internally cooled electrode with that of the conventional 17-G internally cooled electrode. METHODS: A total of 40 (20 for each electrode) and 20 ablation zones (10 for each electrode) were made in extracted bovine livers and in in vivo porcine livers, respectively. Technical parameters, three dimensions [long-axis diameter (Dl), vertical-axis diameter (Dv) and short-axis diameter (Ds)], volume and the circularity (Ds/Dl) of the ablation zone were compared. RESULTS: The total delivered energy was higher in the 15-G group than in the 17-G group in both ex vivo and in vivo studies (8.78 ± 1.06 vs 7.70 ± 0.98 kcal, p = 0.033; 11.20 ± 1.13 vs 8.49 ± 0.35 kcal, p = 0.001, respectively). The three dimensions of the ablation zone had a tendency to be larger in the 15-G group than in the 17-G group in both studies. The ablation volume was larger in the 15-G group than in the 17-G group in both ex vivo and in vivo studies (29.61 ± 7.10 vs 23.86 ± 3.82 cm(3), p = 0.015; 10.26 ± 2.28 vs 7.79 ± 1.68 cm(3), p = 0.028, respectively). The circularity of ablation zone was not significantly different in both the studies. CONCLUSION: The size of ablation zone was larger in the 15-G internally cooled electrode than in the 17-G electrode in both ex vivo and in vivo studies. ADVANCES IN KNOWLEDGE: Radiofrequency ablation of hepatic tumours using 15-G electrode is useful to create larger ablation zones.

E3 ligase activity of RING finger proteins that interact with Hip-2, a human ubiquitin-conjugating enzyme.

To identify proteins that interact with Huntingtin-interacting protein-2 (Hip-2), a ubiquitin-conjugating enzyme, a yeast two-hybrid screen system was used to isolate five positive clones. Sequence analyses showed that, with one exception, all Hip-2-interacting proteins contained the RING finger motifs. The interaction of Hip-2 with RNF2, one of the clones, was further confirmed through in vitro and in vivo experiments. Mutations in the RING domain of RNF2 prevented the clone from binding to Hip-2, an indication that the RING domain is the binding determinant. RNF2 showed a ubiquitin ligase (E3) activity in the presence of Hip-2, suggesting that a subset of RING finger proteins may have roles as E3s.

Wild-type and mutant HIV-1 and HIV-2 Tat proteins expressed in Escherichia coli as fusions with glutathione S-transferase.

Human immunodeficiency virus type 1 (HIV-1) and HIV-2 encode related transcriptional activators known as Tat-1 and Tat-2, respectively, that are required for efficient viral replication. The Tat proteins have been studied extensively, and it appears that their mechanism of action is unique to the primate immunodeficiency viruses or a few distantly related lentiviruses. Here we describe a collection of 24 wild-type and mutant Tat-1 and Tat-2 proteins that are expressed in Escherichia coli as fusions with glutathione S-transferase (GST). The GST-Tat fusions can be used for biochemical studies after simple purification from E. coli lysates in a single step under nondenaturing conditions. The availability of these GST-Tat fusions should be useful to investigators examining biochemical properties of Tat-1 and Tat-2 proteins. E. coli cultures harboring GST-Tat fusions described here are available through the National Institute of Health AIDS Research and Reference Reagent Program.

Construction and characterization of a potent HIV-2 Tat transdominant mutant protein.

The human immunodeficiency virus types 1 and 2 (HIV-1 and HIV-2) Tat proteins Tat-1 and Tat-2 stimulate transcription of the viral long terminal repeat (LTR) sequences and are required for efficient viral replication. A class of mutant Tat proteins, termed "transdominant mutants," has been described that possesses relatively low transactivation activity, yet is able to inhibit the function of wild-type Tat. These mutant proteins contain a nonfunctional TAR RNA-binding domain but apparently retain a functional activation domain. A potential limitation for therapeutic use of transdominant mutants described to date is their low but significant basal level of transactivation for the HIV-1 or HIV-2 LTRs. In order to make an improved transdominant mutant, we have constructed Tat-2 proteins that contain mutations in four contiguous arginines at residues 81 to 84 in the RNA-binding domain. Using purified proteins and in vitro RNA-binding assays, we verified that these mutant Tat-2 proteins are defective for TAR RNA binding. We also verified that these mutant Tat-2 proteins bind to a cellular protein kinase in vitro that we have previously shown to bind specifically to the Tat-1 and Tat-2 activation domain. Using plasmid cotransfection assays, we compared the phenotypes of these mutant Tat-2 proteins with the most potent Tat-1 transdominant mutant described to date. One Tat-2 mutant, named "R81-84A," was found to be equivalent to the Tat-1 mutant in ability to inhibit wild-type Tat transactivation of HIV-1 and HIV-2 LTRs. Moreover, the R81-84A mutant possessed a significantly lower basal level of transactivation than the Tat-1 mutant. The R81-84A Tat-2 mutant is therefore a promising reagent for future development as an anti-HIV agent. Additionally, our results suggest that wild-type Tat-2 transactivation of the HIV-2 LTR is especially sensitive to inhibition by transdominant mutants.

Postsynaptic action mechanism of somatostatin on the membrane excitability in spinal substantia gelatinosa neurons of juvenile rats.

We used tight-seal, whole-cell recording in juvenile rat spinal slices to investigate the action of somatostatin on substantia gelatinosa neurons. Bath application of somatostatin caused a robust and repeatable hyperpolarization or outward current in substantia gelatinosa neurons. Somatostatin inhibited spontaneous action potentials in subpopulation of substantia gelatinosa neurons. The amplitude of dorsal root-evoked excitatory postsynaptic currents and the frequency of spontaneous excitatory postsynaptic currents were not affected by somatostatin. The current induced by somatostatin developed almost instantaneously and did not show any time-dependent inactivation. The current-voltage relationship exhibited inward rectification. The conductance of somatostatin-sensitive current increased with the concentration of external K(+). The reversal potentials in different external K(+) concentrations were close to the K(+) equilibrium potentials. The effect of somatostatin was dose-dependent, with an EC(50) of 113 nM. The somatostatin-sensitive current was blocked by low concentration of extracellular Ba(2+) but not by glibenclamide, an inhibitor of ATP-sensitive K(+) channels. Hyperpolarization-activated cation current in a subpopulation of substantia gelatinosa neurons was not affected by somatostatin. In neurons recorded with an internal solution containing GTPgammaS, somatostatin induced outward current and hyperpolarization that did not reverse on washing. When the spontaneous induction of outward current with GTPgammaS was greatest, somatostatin did not induce any outward currents. Furthermore, intracellular dialysis of GDPbetaS, a G-protein antagonist, abolished the effect of somatostatin. In addition, SST-sensitive neurons were fewer in slices incubated with pertussis toxin than in adjacent control slices incubated without pertussis toxin. These results suggest that somatostatin decreases the postsynaptic membrane excitability of substantia gelatinosa neurons by a pertussis toxin-sensitive G-protein-mediated activation of an inwardly rectifying K(+) conductance.

Mechanism of inhibition of calcium channels in rat nucleus tractus solitarius by neurotransmitters.

1. High-threshold Ca2+ channel currents were measured every 15 s following a 200 ms voltage step from -80 mV to 0 mV in order to study the coupling mechanism between neurotransmitter receptors and Ca2+ channels in neurones acutely isolated from the nucleus tractus solitarius (NTS) of the rat. 2. Application of 30 microM baclofen (GABAB receptor agonist) caused 38.9 +/- 1.2% inhibition of the peak inward Ba2+ current (IBa2+) in most NTS cells tested (n = 85 of 88). Somatostatin, 300 nM, also reduced IBa2+ by 31.3 +/- 1.6% in 53 cells of 82 tested. 3. Activation of mu-opioid-, GABAB- or somatostatin-receptors inhibited both N- and P/Q-type Ca2+ channels. 4. The inhibition of Ca2+ currents by DAMGo (mu-opioid receptor agonist), baclofen and somatostatin was reduced by treatment with pertussis toxin and partially relieved by application of a 50 ms conditioning prepulse to +80 mV. This suggests that a pertussis toxin-sensitive G-protein was involved in the neurotransmitter-mediated action in the observed inhibition of Ca2+ currents. 5. Intracellular loading with an antiserum raised against the amino terminus of Go alpha (GC/2) markedly attenuated the somatostatin-induced inhibition, but did not block the DAMGO- and baclofen-induced inhibition. 6. These findings suggest at least two different pertussis toxin-sensitive G-protein-mediated pathways are involved in receptor-induced inhibition of Ca2+ currents in the NTS.