A high recovery microsampling device based on a microdialysis probe for peptide sampling.

A high recovery microsampling probe based on microdialysis was devised. The new probe showed a high recovery (100%) of peptides in vitro at different perfusion flow rates (0.1-1.0 microl/min). At a high flow rate, 1.0 microl/min, a 10-fold increased in recovery of peptides compared to the conventional microdialysis probe was achieved. A probe made of a low molecular weight cutoff membrane is suitable for filtering off proteins. The new probe can be a useful tool for high recovery of peptides from living tissues.

Clinical significance of the summating potential-action potential ratio and the action potential latency difference for condensation and rarefaction clicks in Meniere's disease.

OBJECTIVES: This study was aimed to elucidate the diagnostic significance of the summating potential (SP)-action potential (AP) ratio and the AP latency difference between condensation and rarefaction clicks (AP con-rar difference) in Meniere's disease. METHODS: The AP and SP were recorded transtympanically in 67 patients with definite Meniere's disease. The SP/AP ratio and the AP con-rar difference were assessed in terms of 1) their interrelationship, 2) their relationship to hearing level, and 3) the rate of occurrence of abnormal values according to the stages of Meniere's disease. RESULTS: No correlation was found between the SP/AP ratio and the AP con-rar difference. Neither the SP/AP ratio in general nor the AP con-rar difference was correlated with the hearing level. However, enhanced values of the SP/AP ratio (0.35 or higher) were moderately correlated with the hearing level (r = 0.51), and their occurrence rate was 55.2%. An increased AP con-rar difference (0.13 ms or longer) was not correlated with the hearing level, and its occurrence rate was 50.2%; it appeared most frequently at stage 3 (p <0.05). CONCLUSIONS: An enhanced SP/AP ratio might not always indicate the presence of endolymphatic hydrops associated with an increase in endolymphatic pressure. An increased AP con-rar difference might reflect the presence of a biased basilar membrane resulting from an increased endolymphatic pressure, and hence it is diagnostically essential to simultaneously evaluate the SP/AP ratio and the AP con-rar difference.

Disposition of protein-bound 3-nitrotyrosine in rat plasma analysed by a novel protocol for HPLC-ECD.

3-Nitrotyrosine (NTyr) is considered as a biomarker of the generation of reactive nitrogen species (RNS). However, it is still difficult to determine its concentration in biological samples. To develop a reliable and high-throughput method, we optimized the conditions for high performance liquid chromatography and electrochemical detection (HPLC-ECD). The best separation of NTyr was achieved using a highly acidic mobile phase (pH 2.5). The concentration of protein-bound NTyr in plasma protein was 593.6 +/- 53.8 fmol/mg in rats treated with lipopolysaccharide (LPS) and 114.4 +/- 27.6 fmol/mg in control. After intravenous administration of in vitro-nitrated plasma protein, NTyr concentration decreased; the half-life was 63.4 +/- 16.8 h. Consistently, protein-bound NTyr concentration in plasma after LPS treatment declined gradually, but was detectable for 1 week. Our protocol is reproducible and suitable for analysing multiple clinical samples to study RNS production in vivo.

Recovery of human compound action potential using a paired-click stimulation paradigm.

The recovery process from adaptation of the compound action potential (CAP) was studied using an equilevel paired click stimulation paradigm in humans with normal hearing. The CAP amplitude to the second click of paired clicks was normalized to the amplitude to the first click. The second-click CAP amplitude recovered as a function of interclick interval (ICI) between the first and the second click of a pair. A regression line fitted to the recovered amplitude data demonstrated the logarithmic function of the ICI. Full recovery times changed from 118 to 278 ms with increasing click intensity. The regression lines for higher click intensities exhibited two different slopes in two ICI ranges: from 3 to 100 and 120 to 300 ms. We suppose that the CAP recovery for ICI <100 ms is attributable to both the relative refractoriness of auditory nerve and the short-term adaptation mechanisms, while, for ICI >100 ms chiefly to the short-term adaptation mechanisms. The recovery process of the second-click CAP slowed with increasing intensity, which is a similar result to that obtained in the animal experiments by Parham et al. The input-output (I-O) curve of the second-click CAP amplitudes exhibited a different slopes above and below 60 dB normal hearing level (nHL). We assume that the mechanisms underlying this characteristic curve pattern differ from those for the I-O curve of the CAP in response to single-click stimuli. We expect that investigating the CAP recovery in pathological ears will provide clinically useful information on cochlear synaptic function.

Quantitative assessment of protein-bound tyrosine nitration in airway secretions from patients with inflammatory airway disease.

Because reactive nitrogen species (RNS) have potent inflammatory activity, they may be involved in the inflammatory process in pulmonary diseases. We recently reported increased numbers of 3-nitrotyrosine immunopositive cells, which are evidences of RNS production, in the sputum of patients with chronic obstructive pulmonary disease (COPD) and patients with asthma compared with healthy subjects. In the present study, we attempted to quantify this protein nitration in the airways by means of high-performance liquid chromatography (HPLC) used together with an electrochemical detection system that we developed. Sputum samples were obtained from 15 stable COPD patients, 9 asthmatic patients and 7 healthy subjects by using hypertonic saline inhalation. The values for the molar ratio of protein-bound 3-nitrotyrosine/tyrosine in patients with asthma (4.31 +/- 1.13 x 10(-6), p < 0.05) and patients with COPD (3.04 +/- 0.36 x 10(-6), p < 0.01) were significantly higher than those in healthy subjects (1.37 +/- 0.19 x 10(-6)). The levels of protein-bound 3-nitrotyrosine in the airways were not significantly different in asthmatic patients and COPD patients. A significant negative correlation was found between values for protein-bound 3-nitrotyrosine/tyrosine and % FEV1 values in patients with COPD (r = -0.53, p < 0.05) but not in patients with asthma. These results suggest that our HPLC-electrochemical method is useful for quantifying RNS production in human airways. More importantly, they show that increased RNS production in the airways seems to contribute in a critical way to the pathogenesis of COPD, and that the effects of RNS in airways may differ in asthma and COPD.

Effects of nimodipine on quinine ototoxicity.

The compound action potential (CAP) in response to a click train stimulus was recorded at the round window of guinea pigs. Administration of quinine hydrochloride (200 mg/kg) significantly elevated the CAP thresholds by 5 to 25 dB (p < .05), and the CAP waveform elicited by the click train stimulus was abnormal. The amplitude of the CAP elicited by the second click was bigger than that elicited by the first click. These changes may be caused by an abnormally broadened N1 response to the first click in the click train. In contrast, CAP waveforms elicited by the second and subsequent clicks appeared normal. After administration of nimodipine (2 mg/kg), the CAP thresholds and waveforms elicited by the click train stimulus were unchanged. Simultaneous administration of both quinine (200 mg/kg) and nimodipine (2 mg/kg) resulted in the same electrophysiological changes as those induced by quinine alone. These results suggest that nimodipine prevents neither the deterioration in the CAP nor the abnormal properties in the response to a click train stimulus.

Effects of nimodipine on salicylate ototoxicity.

The purpose of this study was to examine the effects of nimodipine on salicylate ototoxicity in guinea pigs. The compound action potential (CAP) was recorded at the round window, and the cochlear blood flow (CBF) was measured simultaneously from the lateral wall of the basal turn of the cochlea by laser Doppler flowmetry. After administration of salicylate (100 mg/kg), the CAP thresholds were significantly elevated, by 5 to 20 dB (p < .05), and the CBF was significantly decreased (p < .05). After administration of nimodipine (2 mg/kg), the CAP thresholds were unchanged, but the CBF had increased significantly (p < .05), while systemic blood pressure had decreased significantly (p < .05). Simultaneous administration of both salicylate (100 mg/kg) and nimodipine (2 mg/kg) resulted in significant elevation of the CAP thresholds (p < .05), while the CBF did not decrease. These results suggest that nimodipine prevents the decrease in CBF induced by salicylate, but that nimodipine does not prevent the deterioration in the CAP.

Quinine affects the response properties of compound action potentials elicited by periodic click trains.

The effects of systemically applied quinine on the compound action potential (CAP) were investigated in 5 guinea pigs. A dose of 200 mg/kg body weight of quinine hydrochloride was administered intramuscularly, and CAPs were recorded at the round window before and after administration. The CAP thresholds of the animals were elevated by 5 to 25 dB approximately 30 minutes after administration, and thresholds recovered in some animals during the experimental session. The CAP waveform elicited by the click train stimulus was abnormal after administration of quinine. The amplitude of the CAPs elicited by the second click was larger than that of those elicited by the first click. These changes may be induced by an abnormally broadened N1 response to the first click in the click train following quinine administration. In contrast, the CAP waveforms elicited by the second click and by the following clicks in the click train appeared normal.

Simultaneous determination of glucose and L-lactate in rat brain by an electrochemical in vivo flow-injection system with an on-line microdialysis sampling.

An electrochemical in vivo flow-injection system with an on-line microdialysis sampling is proposed for the simultaneous monitoring of L-lactate and glucose in rat brain. In the first stage of the operation, the dialysate from the microdialysis probe is delivered to a sample loop of the six-way autoinjector by perfusing Ringer's solution for 80 s at 5 microl min(-1). In the second stage, the dialysate collected in the sample loop is automatically injected for 10 s into the flow-injection line. Injected dialysate is split into two streams and two portions pass through two channels with two different immobilized enzyme reactors (glucose oxidase and lactate oxidase immobilized reactors) to produce hydrogen peroxide from glucose and L-lactate in the dialysate. After a subsequent confluence of the streams, produced hydrogen peroxide can be detected amperometrically at a downstream poly(1,2-diaminobenzene) film-coated platinum electrode, without any interference from oxidizable species and proteins present in the dialysate. Because each channel has a different residence time, two peaks are obtained. The first peak corresponds to L-lactate and the second peak to glucose. The peak current is linearly related to the concentrations of L-lactate between 0.2 and 10 mM and glucose between 0.1 and 20 mM. The present method can be successfully applied to the simultaneous in vivo monitoring of L-lactate and glucose in rat brain. The analytical speed is 45 dialysates h(-1).

Cortical potentials evoked by horizontal rotatory stimulation: the effects of angular acceleration.

OBJECTIVE: To examine the potentials evoked by rotatory stimulation, which we consider the optimal means of invoking angular acceleration and which may lead to clinical applications in patients with vertigo. MATERIAL AND METHODS: Horizontal rotatory stimulation using a trapezoidal waveform was performed with 3 different angular accelerations (20, 15 and 10 degrees/s2) used sequentially, with alternating clockwise and counterclockwise stimulation, to normal subjects and a patient with bilateral loss of vestibular function. RESULTS: It is likely that angular accelerations of < 20 degrees/s2 produce minor somatosensory effects. Clear potentials were evoked in 13/18 ears (72%) of the normal subjects with stimulation at an angular acceleration of 15 degrees/s2. No response was recorded in the patient with stimulation at 15 degrees/s2. CONCLUSION: We suggest that an angular acceleration of 15 degrees/s2 is optimal for clinical examinations.

Determination of nitrotyrosine by HPLC-ECD and its application.

3-nitrotyrosine, a product of tyrosine nitration, is a useful indicator of oxidative damage. We modified the previously reported HPLC-electrochemical detection (ECD) method: specifically, a through-type porous carbon electrode was used as a reducing electrode instead of the mercury-gold amalgam electrode, because the response of the latter changes over time. A combination of reverse-phase HPLC and electrochemical detector passed through -800 mV reduction potential and subsequently under +250 mV oxidation potential allows measurement of 3-nitrotyrosine. The detection limit of this assay was less than 10 fmol. In mice to which lipopolysaccharide (LPS) was administered intraperitoneally, plasma 3-nitrotyrosine levels were elevated, corresponding to LPS dosage. These findings suggest that the improved HPLC-ECD method can be used as a specific and sensitive assay of biological 3-nitrotyrosine and can be applied clinically.

Zinc deficiency and tinnitus.

OBJECTIVE: To determine if there is a correlation between serum zinc levels and audiometric performance in tinnitus patients. METHODS: Seventy-three patients participated in this study. Patient's age was restricted to 20-59 years. All patients were examined at the otolaryngology outpatient clinic of the St. Marianna University Toyoko Hospital. The control group consisted of 38 age- and sex-matched healthy volunteers. A blood sample was taken to measure serum zinc levels. Hypozincemia was set at a level of the mean minus one S.D. in the control group. An average hearing sensitivity was calculated as the mean value of hearing thresholds at five frequencies: 250, 500, 1000, 2000, and 4000 Hz. Normal hearing was indicated when the hearing threshold at each of these frequencies was within 20 dB of normal thresholds. RESULTS: There was no significant difference in serum zinc levels between patients with tinnitus and controls. However, patients with tinnitus who had normal hearing had significantly lower serum zinc levels compared to controls. In contrast, no significant difference in serum zinc levels was found between patients with tinnitus who had hearing loss, and controls. A significant correlation between average hearing sensitivity and serum zinc level was observed. CONCLUSIONS: These findings suggest that zinc is involved in the generation of tinnitus, especially in patients whose hearing is relatively normal.

Patency of inferior meatal antrostomy.

OBJECTIVE: To evaluate the patency and effectiveness of inferior meatal antrostomy for the treatment of maxillary lesions. METHODS: Patients suffering from moderate-to-severe chronic sinusitis who underwent inferior meatal antrostomies at St. Marianna University Toyoko Hospital and followed up for at least 1 year after surgery were selected. Thirty sides of 27 patients were analyzed. Postoperative mucociliary function was examined in seven sides of seven patients. RESULTS: No closure was observed in all 30 procedures. Out of 30 antra, 26 (86.7%) had normal antral mucosa. Recurrent mucosal lesions inside the maxillary sinus were successfully treated through the inferior meatal window. No cases showed circular flow on the india ink test. All sinuses exhibited a discharge from the middle meatal window and three out of seven sinuses exhibited a discharge from the inferior meatal window. CONCLUSIONS: Although long-term follow-up is required to determine the patency of inferior meatal antrostomy, initial results are encouraging. The inferior meatal window provides both intra- and postoperative benefits.

Neurotological findings in a patient with narrow internal auditory canal: a case report.

We report neurotological findings in a patient with unilateral narrow internal auditory canal, as confirmed by computed tomography. The patient presented no auditory brainstem response on the affected side. Vestibular tests including vestibular-evoked myogenic potentials (VEMP) and caloric test revealed normal function of both inferior and superior vestibular neural pathways.

A case of meningoencephalic herniation of the temporal bone.

A case of cerebrospinal fluid leakage from a meningoencephalic herniation of the temporal bone is presented. Computed tomography and magnetic resonance imaging were useful for a diagnosis of meningoencephalic herniation. After surgical treatment, no recurrence was observed during a 6-month follow-up period.

Subcutaneous emphysema secondary to tonsillectomy: a case report.

We report a patient in whom subcutaneous emphysema developed shortly after a tonsillectomy. A 55-year-old female with a chronic tonsillitis underwent a tonsillectomy, and about 8 h after surgery complained of swelling to the left side of the face and neck. There was crepitus and local tenderness in the left side of the neck. A CT scan revealed subcutaneous emphysema. The mechanisms underlying emphysema after tonsillectomy are discussed.

Changes in CAP adaptation in experimentally induced endolymphatic hydrops.

OBJECTIVE: To determine the effect of experimentally induced endolymphatic hydrops (ELHs) on compound action potential (CAP) adaptation. METHODS: The CAP was recorded 1 month after surgical treatment to produce ELH. The CAP threshold in response to click stimuli showed no significant differences between the hydropic and control groups. The CAP was elicited from the retroauricular-exposed round window using a silver ball electrode. Click trains consisting of 10 clicks with an interclick interval (Deltat) of either 4 or 8 ms were delivered through a loudspeaker placed in front of the animals' ears. The patterns of adaptation to these click trains were evaluated as functions of Deltat and intensity level. RESULTS: An abnormal pattern of CAP adaptation was observed in hydropic ears. CONCLUSIONS: These findings suggest that early ELH influences CAP adaptation.

A case report of difficult diagnosis in the patient with advanced laryngeal tuberculosis.

We report a patient with severe laryngeal tuberculosis (LTB) involving thyroid cartilage and combined with whole-bone metastasis. A 57-year-old male had presented only with hoarseness. Radiological findings were indicative of suspected metastasis from a malignant tumor. However, tuberculosis was considered by histopathological findings, and so sputum samples were tested for acid-fast bacilli and purified protein derivatives of tuberculin in order to detect the presence of LTB. A polymerase chain reaction confirmed the diagnosis. Anti-tuberculous medications were effective in resolving the hoarseness, and the removal of the mass in the right wing of thyroid cartilage was confirmed by computed tomography (CT).

The recovery from AP adaptation in sensorineural hearing loss.

CONCLUSIONS: An abnormally slower action potential (AP) recovery from adaptation (decreased recovery, dR) was characteristically detected in many ears with sensorineural hearing loss (SNHL) but not Meniere's disease and idiopathic sudden SNHL. We assumed that this abnormal AP recovery from adaptation was attributed to an imbalance in the distributions of auditory neurons with high and low spontaneous firing rates. The significant difference of initial hearing level between dR and normal AP recovery groups (nR) was assumed to partially result from AP recovery being determined by the inner hair cell synapse, and not from outer hair cells. OBJECTIVE: This study aimed to detect the AP recovery pattern in SNHL. METHODS: Electrocochleography (ECochG) was performed transtympanically in 30 patients with SNHL. AP recovery was measured by a paired click stimulation paradigm as a function of inter-click intervals from 5 to 100 ms. RESULTS: The high prevalence of dR (9 of 30 ears) appears to be a characteristic ECochG finding in SNHL. Initial hearing level differed significantly between dR and nR groups.

Electrocochleography in Meniere's disease: recovery from adaptation of the auditory action potential.

CONCLUSIONS: High prevalence rate of an enhanced summating potential (SP)/action potential (AP) ratio (≥0.35) is considered to result largely from the imbalances of Ca(2+) homeostasis in the endolymphatic space. An increase in Ca(2+) is thought to augment the SP. Furthermore, a greater inflow of Ca(2+) into inner hair cells is supposed to influence the disturbed release of neurotransmitter, which may induce pathological functioning of inner hair cell synapses. We speculate that such abnormal synaptic functioning is responsible for the development of abnormally increased AP recovery from adaptation (iR). OBJECTIVES: This study aimed to record the AP recovery from adaptation (AP recovery) and to investigate its clinical significance in association with other electrocochleography (ECochG) findings in Meniere's disease. METHODS: ECochG was performed transtympanically in 27 patients with Meniere's disease. AP recovery was measured by a paired-click stimulation paradigm as a function of interclick intervals from 5 to 100 ms. RESULTS: High prevalence of SP/AP ratio was observed in 27 patients. iR, suggesting abnormal functioning of inner hair cell synapse, was in many cases combined with an enhanced SP/AP ratio.