Muscarinic receptor-mediated excitation of rat intracardiac ganglion neurons.

Modulation of the membrane excitability of rat parasympathetic intracardiac ganglion neurons by muscarinic receptors was studied using an amphotericin B-perforated patch-clamp recording configuration. Activation of muscarinic receptors by oxotremorine-M (OxoM) depolarized the membrane, accompanied by repetitive action potentials. OxoM evoked inward currents under voltage-clamp conditions at a holding potential of -60 mV. Removal of extracellular Ca(2+) markedly increased the OxoM-induced current (IOxoM). The inward IOxoM in the absence of extracellular Ca(2+) was fully inhibited by removal of extracellular Na(+), indicating the involvement of non-selective cation channels. The IOxoM was inhibited by organic cation channel antagonists including SKF-96365 and ML-204. The IOxoM was antagonized by muscarinic receptor antagonists with the following potency: 4-DAMP > pirenzepine = darifenacin > methoctramine. Muscarinic toxin 7 (MT-7), a highly selective inhibitor for M1 receptor, produced partial inhibition of the IOxoM. In the presence of MT-7, concentration-inhibition curve of the M3-preferring antagonist darifenacin was shifted to the left. These results suggest the contribution of M1 and M3 receptors to the OxoM response. The IOxoM was inhibited by U-73122, a phospholipase C inhibitor. The membrane-permeable IP3 receptor blocker xestospongin C also inhibited the IOxoM. Furthermore, pretreatment with thapsigargin and BAPTA-AM inhibited the IOxoM, while KN-62, a blocker of Ca(2+)/calmodulin-dependent protein kinase II, had no effect. These results suggest that the activation mechanism involves a PLC pathway, release of Ca(2+) from intracellular Ca(2+) stores and calmodulin. The cation channels activated by muscarinic receptors may play an important role in neuronal membrane depolarization in rat intracardiac ganglion neurons.

Is recall of dreaming during anesthesia a sign of occurrence of postoperative nausea and vomiting?

We examined the relationships between recall of dreaming during anesthesia and postoperative nausea and vomiting (PONV). We found a relationship between PONV within 24 h and age <50 years, use of postoperative epidural analgesia with morphine, and female gender. We also found a relationship between PONV lasting more than 24 h and dream recall. As serotonin plays an important role for both inducing PONV and dream recall, results of the present study may suggest a possible relationship between dream recall and PONV.

The supine-to-prone position change induces modification of endotracheal tube cuff pressure accompanied by tube displacement.

STUDY OBJECTIVES: To determine whether the supine-to-prone position change displaced the endotracheal tube (ETT) and, if so, whether the displacement related to this change modified ETT cuff pressure. DESIGN: Prospective study. SETTING: Operating room of a university hospital. PATIENTS: 132 intubated, adult, ASA physical status 1, 2, and 3 patients undergoing lumbar spine surgery. INTERVENTIONS AND MEASUREMENTS: After induction of anesthesia, each patient's trachea was intubated. The insertion depth of each ETT was 23 cm for men and 21 cm for women at the upper incisors. In the supine position and after the supine-to-prone position change with the head rotated to the right, the length from the carina to ETT tip and ETT cuff pressure were measured. MAIN RESULTS: After the supine-to-prone position change, 91.7% patients had ETT tube displacement. Of these, 48% of patients' ETT moved ≥ 10 mm, whereas 86.3% of patients had changes in tube cuff pressure. There was a slight but significant correlation between ETT movement and change in cuff pressure. Depending on the position change, ETT cuff pressure decreased and the ETT tended to withdraw. CONCLUSIONS: After the supine-to-prone position change, patients had ETT tube displacement. Such ETT movement may be accompanied by a decrease in cuff pressure.

Overcoming regulatory T-cell suppression by a lyophilized preparation of Streptococcus pyogenes.

Cancer vaccines have yet to yield clinical benefit, despite the measurable induction of humoral and cellular immune responses. As immunosuppression by CD4(+) CD25(+) regulatory T (Treg) cells has been linked to the failure of cancer immunotherapy, blocking suppression is therefore critical for successful clinical strategies. Here, we addressed whether a lyophilized preparation of Streptococcus pyogenes (OK-432), which stimulates Toll-like receptors, could overcome Treg-cell suppression of CD4(+) T-cell responses in vitro and in vivo. OK-432 significantly enhanced in vitro proliferation of CD4(+) effector T cells by blocking Treg-cell suppression and this blocking effect depended on IL-12 derived from antigen-presenting cells. Direct administration of OK-432 into tumor-associated exudate fluids resulted in a reduction of the frequency and suppressive function of CD4(+) CD25(+) Foxp3(+) Treg cells. Furthermore, when OK-432 was used as an adjuvant of vaccination with HER2 and NY-ESO-1 for esophageal cancer patients, NY-ESO-1-specific CD4(+) T-cell precursors were activated, and NY-ESO-1-specific CD4(+) T cells were detected within the effector/memory T-cell population. CD4(+) T-cell clones from these patients had high-affinity TCRs and recognized naturally processed NY-ESO-1 protein presented by dendritic cells. OK-432 therefore inhibits Treg-cell function and contributes to the activation of high-avidity tumor antigen-specific naive T-cell precursors.

Antibody responses against NY-ESO-1 and HER2 antigens in patients vaccinated with combinations of cholesteryl pullulan (CHP)-NY-ESO-1 and CHP-HER2 with OK-432.

Combination vaccines of the NY-ESO-1 protein complexed with cholesteryl pullulan (CHP), CHP-NY-ESO-1, and the truncated 146HER2 protein with CHP, CHP-HER2, were subcutaneously administered with the immuno-adjuvant OK-432 to eight esophageal cancer patients. Vaccination was well-tolerated. NY-ESO-1- and HER2-specific antibody responses were analyzed using the patients' sera and samples from previous single CHP-NY-ESO-1 or CHP-HER2 vaccine trial. The responses to NY-ESO-1 in the combination vaccine study were comparable to the single vaccine. For responses to HER2, there were fewer antibody responses in the combination vaccines. Although there were marked individual variations in the antibody responses to the NY-ESO-1 and HER2 antigens, the reaction patterns to these antigens were comparable within each patient. Antibodies to OK-432 were not augmented. Protein cancer vaccines targeting multiple antigens are feasible.

Regulatory T cell-resistant CD8+ T cells induced by glucocorticoid-induced tumor necrosis factor receptor signaling.

We previously found that a Salmonella typhimurium vector engineered to secrete soluble tumor antigen induces CD4(+) T cells resistant to CD4(+)CD25(+) regulatory T cells (Treg) and that glucocorticoid-induced tumor necrosis factor receptor family-related gene (GITR) signal is involved in the development of this resistance. In this study, we address the potential of incorporating GITR ligand (GITRL) as a way to augment the immunogenicity of cancer vaccines. BALB/c mice were immunized by gene gun with plasmids encoding the mutated extracellular signal-regulated kinase 2 (mERK) with or without plasmids encoding mouse GITRL. Coadministration with GITRL during primary and secondary immunization enhanced the induction of mERK-specific CD8(+) T cells. Antibody depletion and minigene analysis suggested that GITRL directly activated CTL epitope-specific CD8(+) T cells independently of CD4(+) T cells. Immunization with plasmids encoding a CTL epitope and GITRL resulted in strong tumor inhibition in a CD8(+) T cell-dependent manner. Furthermore, CTL epitope-specific CD8(+) T cells induced by immunization with plasmids encoding CTL epitope coadministered with GITRL were refractory to suppression by CD4(+)CD25(+) Tregs compared with CD8(+) T cells induced without GITR signaling. We propose that coadministration of GITR signaling agents with tumor antigens constitutes a promising novel strategy for cancer vaccine development.

Humoral immune responses in patients vaccinated with 1-146 HER2 protein complexed with cholesteryl pullulan nanogel.

The CHP-HER2 vaccine, comprising truncated 146HER2 protein complexed with nanogels of cholesteryl pullulan (CHP), is a novel protein antigen vaccine that elicits 146HER2-specific CD8(+) and CD4(+) T-cell immune responses in patients with HER2-expressing tumors. We analyzed the humoral responses in patients vaccinated with CHP-HER2 and those with CHP-HER2 plus granulocyte-macrophage colony-stimulating factor (GM-CSF). The vaccine was injected subcutaneously at a dose of 300 microg protein. Nine patients received the vaccine alone over the first four injections, followed by CHP-HER2 with GM-CSF or OK-432, whereas six received CHP-HER2 plus GM-CSF from the first cycle. 146HER2-specific IgG antibodies were induced in 14 patients, who were negative at baseline. The antibodies became detectable after the second or third vaccination and reached plateau levels after the third or fourth cycle in patients vaccinated with CHP-HER2 plus GM-CSF. In contrast, the antibodies appeared only after the third to sixth vaccination and the plateau appeared after the fourth to eighth cycle in patients vaccinated with the CHP-HER2 vaccine alone over the first four cycles. The antibodies induced by the vaccine were not reactive with HER2 antigen expressed on the cell surface in any of the patients. Epitope analysis using overlapping peptides revealed a single region in the 146HER2 protein, amino acids 127-146, in eight patients who were initially vaccinated with CHP-HER2 alone. Similarly, the same HER2 region was recognized dominantly in patients vaccinated with GM-CSF. Our results indicate that CHP-HER2 induced HER2-specific humoral responses in patients with HER2-expressing tumors and that GM-CSF seems to accelerate the responses.