Activation of presynaptic 5-HT3 receptors facilitates glutamatergic synaptic inputs to area postrema neurons in rat brain slices.

Whole-cell voltage-clamp recordings were performed to investigate the serotonergic modulation of neurotransmitter release onto rat area postrema neurons in vitro. The bath application of serotonin (5-HT; 50 microM) or phenylbiguanide (PBA; 50 microM), a potent 5-HT3 receptor agonist, increased the frequency of spontaneous excitatory postsynaptic currents (sEPSCs) or miniature EPSCs (mEPSCs) in 35 of 83 neurons (42%). These increases occurred in all electrophysiological cell classes. No cells exhibited a decrease in EPSC frequency. The majority of responding cells showed no inward currents during the application of serotonergic agonists (n = 34/35). However, the amplitude of mEPSCs was increased in 11/11 cells with 5-HT or 3/11 cells with PBA. ICS-205,930, a potent 5-HT3 receptor antagonist, markedly suppressed the 5-HT-induced facilitation of sEPSCs (n = 5) or mEPSCs (n = 5). An increase in the frequency of mEPSCs after PBA exposure was found, even with media containing Cd2+ (50 microM) or zero Ca2+. mEPSCs and evoked EPSCs were completely blocked in media containing the non-NMDA ionotropic receptor antagonist, CNQX (10 microM), indicating that EPSCs were glutamate events. These results suggest that glutamate release is increased in the area postrema by presynaptic 5-HT3 receptor activation. Furthermore, we present evidence that 5-HT3 receptor activation may be able to directly release glutamate from terminals, bypassing a requirement for voltage-dependent calcium entry into terminals. Such a mechanism may contribute to the chemosensitive function of area postrema neurons.

Effects of amiloride on gustatory neural responses to salts in the frog.

In frogs, the glossopharyngeal nerve (GL) innervates taste receptors on almost the entire tongue. The mandibular branch (MBF) and palatine branch (PN) of the facial nerve innervate taste receptors on a very small area at the base of the tongue and on the palate, respectively. In the present study, effects of amiloride, an epithelial sodium channel blocker, on the tonic responses of the GL, MBF and PN in frogs to NaCl, LiCl, KCl and CaCl(2) were investigated. In three nerves, amiloride at 0.5 mM, a relatively high concentration, did not affect the responses to 0.15 (concentration just above threshold)-0.5 M NaCl, 0.5 M LiCl and 0.3 M KCl, whereas it almost completely inhibited the response to 1.0 mM CaCl(2). Amiloride may exert an inhibitory action on the response to CaCl(2) by a competitive antagonism between Ca(2+) and a monovalent cation of amiloride, because the response to Ca(2+) is competitively inhibited by other cations such as Na(+) and Mg(2+). The lack of inhibitory effect of amiloride on the responses in the GL, MBF and PN to NaCl suggests that amiloride-sensitive sodium channels in the apical membrane of taste receptor cells are not involved in sodium taste transduction in frogs.

Determination of the antigen/epitope that is recognized by human monoclonal antibody CLN-IgG.

Ever since the development of human monoclonal antibody CLN-IgG in 1982, we anticipated the identification of the antigen that is recognized by this antibody. Despite its scarce expression on the cell surface, susceptibility to proteolytic enzymes and adherence to experimental equipment, we finally succeeded in determining the antigen moiety that is recognized by this antibody by means of CLN-IgG conjugated column affinity chromatography, two-dimensional electrophoresis, MALDI-TOF/MS and use of glioblastoma cell line U-251MG. The antigen was found to be vimentin, a cytoskeletal protein, and we succeeded in determining a 79 amino acids sequence of the epitope which turned out to comprise a part of the c2 (coil 2 of the central rod) domain of vimentin.

Role of parabrachial nucleus in submandibular salivary secretion induced by bitter taste stimulation in rats.

When rats lick a bitter taste solution such as quinine-hydrochloride, they secrete profuse amounts of saliva. The salivation has a higher flow rate than that induced by other qualities of taste stimulation: sweet, salty, and sour. The present study is aimed to clarify the neural mechanism of the quinine-evoked salivation by means of behavioral, neuroanatomical, and electrophysiological experiments. Behaviorally, submandibular salivary secretion and rejection behavior (gaping) were observed in normal rats, as well as in rats chronically decerebrated at the precollicular level. In chronically decerebrate rats, these quinine-evoked reactions were strongly suppressed by destruction of the medial part of the parabrachial nucleus, including the so-called taste area, and ventral part of the parabrachial nucleus, including the pontine reticular formation. Neuroanatomical study using a retrograde tracer, Fluoro-gold, revealed that the neurons sending their axons to the superior salivatory nucleus, parasympathetic secretory center, were located mainly in the pontine reticular formation ventral to the parabrachial nucleus, not in the parabrachial taste area. Extracellular neural activity was recorded from the parabrachial region in decerebrate rats, and responsiveness to taste stimulation, jaw movements, and electrical stimulation of the superior salivatory nucleus was examined. Neurons responsive to both taste stimulation and antidromic stimulation of the superior salivatory nucleus were found in the pontine reticular formation ventral to the parabrachial nucleus, which responded well to quinine and HCl taste stimuli. Neurons in the parabrachial taste area could respond to four qualities of taste stimulation, but not to antidromic stimulation of the salivary center. These results suggest that aversive taste information from the parabrachial taste area reaches the salivary secretory center via the reticular formation ventral to the parabrachial nucleus.

Intravenous anesthetics inhibit nonadrenergic noncholinergic lower esophageal sphincter relaxation via nitric oxide-cyclic guanosine monophosphate pathway modulation in rabbits.

BACKGROUND: Nonadrenergic noncholinergic (NANC) nerves have important roles in the regulation of the lower esophageal sphincter (LES) motility and function. The effects of thiopental, ketamine, and midazolam on NANC LES relaxation were investigated. METHODS: The isometric tension of circular muscle strips from Japanese White rabbits was examined. The NANC relaxation was induced by KCl (30 mM) in the presence of atropine (3 x 10(-6) M) and guanethidine (3 x 10(-6) M). The modifications of the NANC and sodium nitroprusside (SNP; 10(-5) M)-induced relaxation by the anesthetics were examined. The content of 3',5'-cyclic guanosine monophosphate (cGMP) was measured by radioimmunoassay. RESULTS: The KCl-induced relaxation was abolished by pretreating with tetrodotoxin (10(-6) M). The NANC relaxation was inhibited in the presence of N(G)-nitro-L-arginine (L-NNA; 3 x 10(-5) M), methylene blue (10(-6) M), apamin (10(-7) M), and glibenclamide (10(-5) M). The SNP-induced relaxation was inhibited by methylene blue but was not affected by tetrodotoxin, L-NNA, apamin, or glibenclamide. Ketamine (EC50 = 8.8 x 10(-5) M) and midazolam (EC50 = 4.8 x 10(-6) M) suppressed the NANC response in a concentration-dependent manner, leaving SNP-induced response unchanged. Thiopental altered neither of the relaxations. cGMP content was decreased in the presence of ketamine and midazolam. CONCLUSION: The NANC relaxation was mediated by nitric oxide and by low-conductance calcium- and adenosine triphosphate-sensitive potassium channels of smooth muscle. The modulation of the nitric oxide-cGMP pathway was related, at least in part, to the inhibitory actions of ketamine and midazolam on the NANC LES relaxation.

Salt taste responses of the IXth nerve in Sprague-Dawley rats: lack of sensitivity to amiloride.

To explore characteristics of the salt taste function of taste receptor cells located on the posterior tongue, we recorded electrophysiological responses from the whole glossopharyngeal nerve in Sprague-Dawley (SD) rats. For all salts, relative response magnitudes increased with increased stimulus concentrations (0.2-2.0 M) of NH4+, K+, and Na+ salts. The order of effectiveness of stimulation for Cl- salts was NH4Cl > KCl > NaCl. For sodium salts, relative response magnitudes were anion dependent. Sodium salts with small anions (NaCl, NaSCN, and NaNO3) had a much stronger stimulating effect than sodium salts with large anion groups (Na2SO4, C2H3O2Na, and C6H11O7Na). The responses of the glossopharyngeal nerve to the Na+ salts of NaCl, C2H3O2Na, and C6H11O7Na were not inhibited by the lingual application of the epithelial sodium transport blocker amiloride. This is in contrast to large amiloride sensitivity of the chorda tympani nerve. Amiloride also failed to inhibit the responses to K+ salts (KCl and KC2H3O2) and to NH4Cl. These results demonstrate that taste receptors innervated by the glossopharyngeal nerve in SD rats lack amiloride sensitivity as observed in the glossopharyngeal nerve of spontaneously hypertensive and Wistar-Kyoto rats. Furthermore, the difference between the small-anion group and the large-anion group of Na+ salts in their effectiveness to produce responses in the glossopharyngeal nerve parallels the effects noted for the anion dependence in the portion of the taste response resistant to amiloride in the chorda tympani nerve. Sodium salts with the smaller anion produced the larger responses in both glossopharyngeal and chorda tympani nerves after amiloride.

Effects of a bite-raising splint on the duration of the chewing cycle and the EMG activities of masticatory muscles during chewing in freely moving rabbits.

Metal bite-raising splints of 0.5 mm thickness were attached to the upper molar teeth on both sides of the jaw in rabbits. The effects of these splints on masticatory behaviour during the chewing of soft food (bread) by freely moving rabbits were investigated. We recorded electromyograms (EMGs) of the masseter and digastric muscles. The animals exhibited prolongation of the chewing cycle, decreased EMG activity of the masseter muscle and increased EMG activity of the digastric muscle during chewing after introduction of the bite-raising splints. The effects of the splints on the activities of masticatory muscles were abolished by bilateral sectioning of the maxillary and inferior alveolar nerves. It seems likely that afferents from oral sensory receptors were responsible for the changes in masticatory behaviour after the introduction of the occlusal splint.

Enhancing effect of nickel ions on the response to magnesium ions of single fibers of the frog glossopharyngeal nerve: competitive inhibition by calcium ions of the nickel-enhanced response to magnesium ions.

Single fibers of the frog glossopharyngeal nerve respond to MgCl2 at concentrations exceeding 10 mM. NiCl2 at 1 mM enhanced the Mg2+ response. CaCl2 at 0.5-2 mM induced an inhibition of the Ni(2+)-enhanced response to Mg2+ ions. A quantitative explanation for these results is provided by the hypothesis that Ni2+ ions secondarily affect a magnesium receptor (designated X*Mg) that is responsible for the Mg2+ response and that Ca2+ ions inhibit the Ni(2+)-enhanced response to Mg2+ ions by competing with Mg2+ ions for X*Mg. Double-reciprocal plots of the experimental data indicate that Ni2+ ions do not affect the affinities of X*Mg for both Mg2+ ions (agonist) and Ca2+ ions (competitive antagonist) appreciably, and that Ni2+ ions at 1 mM enhanced the maximal response to Mg2+ ions by 270%. It appears that a magnesium receptor interacts with an Ni(2+)-binding element that is affected by Ni2+ ions and, thus, Ni2+ ions can induce an enhancement of the Mg2+ response.

A quantitative study of the enhancing effect of nickel ions on the taste response to sodium ions of single fibers of the frog glossopharyngeal nerve: competitive inhibition by calcium ions of the nickel-enhanced response to sodium ions.

Single water fibers of the frog glossopharyngeal nerve respond to relatively high concentrations of NaCl ( > 80 mM). NiCl2 at 1 mM enhanced the Na+ response and reduced the threshold concentration for NaCl to 20 mM. CaCl2 at 0.5-1 mM induced an inhibition of the Ni2+ -enhanced response to Na+ ions. A quantitative explanations for these results is provided by the hypothesis that Ni2+ ions secondarily affect a sodium receptor or channel (designated XNa*) that is responsible for the Na+ response and that Ca2+ ions inhibit the Ni2+ -enhanced response to Na+ ions by competing with Na+ ions for XNa*. Double-reciprocal plots of the experimental data indicate that the affinity of XNa* for both Na+ ions (agonist) and Ca2+ ions (competitive antagonist) in the presence of 1 mM NiCl2 was five times higher than the previously reported values obtained in the absence of NiCl2 (Kitada, 1991). Ni2+ ions at 1 mM enhanced the maximal response to Na+ ions by 190%. It appears that a sodium receptor (or channel) interacts with a Ni2+ -binding element that is affected by Ni2+ ions and, thus, Ni2+ ions can induce both an increase in the affinity of the sodium receptor for the respective cations and an enhancement of the Na+ response.

Iatrogenic acute spinal epidural abscess with septic meningitis: MR findings.

A contaminated catheter used in epidural anesthesia in a 71-year-old female produced acute epidural abscess and septic meningitis. Methicillin-resistant Staphylococcus aureus (MRSA) was detected in a culture of the epidural pus. Both T1- and T2-weighted MR images showed low intensity mass lesion compressing the thecal sac behind the vertebral body L3. The low intensity lesion was probably pus with gas component. In these low intensity lesions in MR findings with gas component, MR was superior to myelography because it visualized both the degree of compression to the thecal sac and extension of the lesion in all directions.

Antiscorbutic activity of L-ascorbic acid 2-glucoside and its availability as a vitamin C supplement in normal rats and guinea pigs.

Bioavailability of a newly-synthesized and chemically-stable 2-O-alpha-D-glucopyranosyl-L-ascorbic acid (AA-2G) as a vitamin C supplement was investigated in rats and guinea pigs. Oral administration of AA-2G to the animals resulted in an increase of serum ascorbic acid (AA) levels. However, in these sera the intact form was not detectable by the high performance liquid chromatography (HPLC) method, indicating its hydrolysis through the process of absorption. After an intravenous injection of AA-2G, the intact form diminished rapidly from the serum, followed by prolonged and marked elevation of serum AA levels. Various tissue homogenates from rats and guinea pigs were examined for their releasing activity of AA from AA-2G. High activity was observed in kidney, small intestine and serum of rats and in small intestine and kidney of guinea pigs. These hydrolytic activities were completely inhibited by castanospermine, a specific alpha-glucosidase inhibitor, suggesting the participation of alpha-glucosidase in the in vivo hydrolysis of AA-2G. AA-2G was found to exhibit obvious therapeutic effect in scorbutic guinea pigs by its repeated oral administrations. These results indicate that AA-2G is a readily available source of vitamin C activity in vivo.

Development of a sensitive enzyme immunoassay for OPC-7251, a novel antimicrobial agent for percutaneous application.

A sensitive enzyme immunoassay for OPC-7251, a novel pyridone carboxylic acid antimicrobial agent, was developed and applied for the determination of human plasma levels. OPC-7251 was coupled to bovine serum albumin through a formation of N-hydroxysuccinimide ester. By immunization of rabbits, highly specific antiserum was raised. Using the antiserum and beta-D-galactosidase-labeled hapten, the homologous assay system allowed the detection of 2 pg of this compound. Plasma samples were precisely analyzed down to the minimum value of 200 pg/ml after heat treatment. The system was further validated by the recovery test and correlation with the HPLC analyses. Percutaneous application of 10 g of 1% OPC-7251 cream to healthy volunteers resulted in the peak plasma value of 1.6 ng/ml about 8 hours after dosing, indicating extremely low absorption efficiency through a transdermal system.

Follicular cyst derived from hair matrix and outer root sheath.

A new follicular cyst was reported. The lower part of the cyst wall was composed of both basophilic and shadow cells as seen in pilomatricoma, whereas the upper part of the wall consisted of clear cells. Our case apparently derives from hair matrix and outer root sheath.

[Erythema action spectrum in xeroderma pigmentosum group A patients].

Twenty patients with xeroderma pigmentosum group A (XP-A) and 30 healthy controls were studied to determine the energy requirements for UV-erythema with monochromatic light between 255 and 340 nm. As test sites, unexposed skin on the upper back was selected for irradiation using a prism monochromator and a 1 kW xenon arc source. The minimal erythema doses (MEDs) were determined at each erythemal response peak, at 24 hours post irradiation in normal controls and at 72 hours in XP-A patients, respectively. Compared with normal subjects, XP-A patients had extremely low MEDs at all wavelengths tested. The erythema action spectrum showed 2 peaks at 265 and 290 nm in XP-A patients and the greatest effectiveness at 290 nm. These results indicate that XP-A patients should be protected strongly against not only UVB but also UVA, because the photo-hypersensitivity in XP-A patients has been extended from the UVB to UVA region.

Fetal case of xeroderma pigmentosum--first report of an autopsy case.

Fetal autopsy case of xeroderma pigmentosum was reported. This male fetus of 24 gestational weeks was prenatally diagnosed as xeroderma pigmentosum by detecting the DNA-repair defect of the amniotic fluid cells. Autopsy revealed not only maldevelopment of the fetus (stillbirth) in general for the standard, but also showed slight developmental retardation of various organs including kidneys and lungs, which could be examined by microscopic analysis. It was suggested that abnormality of the somatic organs began to appear at the fetal stage in this case of xeroderma pigmentosum.