Cloning and characterization of decaprenyl diphosphate synthase from three different fungi.

Coenzyme Q (CoQ) is composed of a benzoquinone moiety and an isoprenoid side chain of varying lengths. The length of the side chain is controlled by polyprenyl diphosphate synthase. In this study, dps1 genes encoding decaprenyl diphosphate synthase were cloned from three fungi: Bulleromyces albus, Saitoella complicata, and Rhodotorula minuta. The predicted Dps1 proteins contained seven conserved domains found in typical polyprenyl diphosphate synthases and were 528, 440, and 537 amino acids in length in B. albus, S. complicata, and R. minuta, respectively. Escherichia coli expressing the fungal dps1 genes produced CoQ10 in addition to endogenous CoQ8. Two of the three fungal dps1 genes (from S. complicata and R. minuta) were able to replace the function of ispB in an E. coli mutant strain. In vitro enzymatic activities were also detected in recombinant strains. The three dps1 genes were able to complement a Schizosaccharomyces pombe dps1, dlp1 double mutant. Recombinant S. pombe produced mainly CoQ10, indicating that the introduced genes were independently functional and did not require dlp1. The cloning of dps1 genes from various fungi has the potential to enhance production of CoQ10 in other organisms.

[Consideration on molecular imaging technology as a tool for drug research and development].

Molecular imaging technology such as positron emission tomography (PET) and magnetic resonance imaging (MRI) are known as powerful tools for clinical diagnosis in neurology, oncology and so on. As applications to new drug research and development, there are three methodologies which are PK (Pharmacokinetics study), PD (Pharmacodynamic study), and efficacy study. When we use these methodologies for the drug research, we must consider construction of technological environment (tracer, animal model, imaging analysis software, and clinical database) and regulatory environment for GMP (Good Manufacturing Practice) and GCP (Good Clinical Practice) level. Additionally, concept of microdosing and exploratory clinical study was proposed in western countries and the guidance on microdosing study was also announced by Health, Labor and Welfare Ministry on June 3rd 2008. However they may be still in learning phase, we must meet with complexity, high cost, and indigestion. To promote molecular imaging technology into the drug research, integration of the scientists between academia and industry is important because it needs much type of the advanced technologies and skills.

Partial volume effect-corrected FDG PET and grey matter volume loss in patients with mild Alzheimer's disease.

PURPOSE: Although( 18)F-fluorodeoxyglucose (FDG) PET is an established imaging technique to assess brain glucose utilisation, accurate measurement of tracer concentration is confounded by the presence of partial volume effect (PVE) due to the limited spatial resolution of PET, which is particularly true in atrophic brains such as those encountered in patients with Alzheimer's disease (AD). Our aim was to investigate the effects of PVE correction on FDG PET in conjunction with voxel-based morphometry (VBM) in patients with mild AD. METHODS: Thirty-nine AD patients and 73 controls underwent FDG PET and MRI. The PVE-corrected grey matter PET images were obtained using an MRI-based three-compartment method. Additionally, the results of PET were compared with grey matter loss detected by VBM. RESULTS: Before PVE correction, reduced FDG uptake was observed in posterior cingulate gyri (PCG) and parieto-temporal lobes (PTL) in AD patients, which persisted after PVE correction. Notably, PVE correction revealed relatively preserved FDG uptake in hippocampal areas, despite the grey matter loss in medial temporal lobe (MTL) revealed by VBM. CONCLUSION: FDG uptake in PCG and PTL is reduced in AD regardless of whether or not PVE correction is applied, supporting the notion that the reduced FDG uptake in these areas is not the result of atrophy. Furthermore, FDG uptake by grey matter tissue in the MTL, including hippocampal areas, is relatively preserved, suggesting that compensatory mechanisms may play a role in patients with mild AD.

Evaluation of (+)-p-[11C]methylvesamicol for mapping sigma1 receptors: a comparison with [11C]SA4503.

Vesamicol is a leading compound for positron emission tomography (PET) and single photon emission computed tomography (SPECT) tracers for mapping the vesicular acetylcholine transporter (VAChT). Recently, we found that (+)-p-methylvesamicol ((+)-PMV) has low affinity for VAChT (K(i)=199 nM), but has moderate to high affinity for sigma receptors: K(i)=3.0 nM for sigma1 and K(i)=40.7 nM for sigma2, and that sigma1-selective SA4503 (K(i)=4.4 nM for sigma1 and K(i)=242 nM for sigma2) has moderate affinity for VAChT (K(i)=50.2 nM). In the present study, we examined the potential of (+)-[11C]PMV as a PET radioligand for mapping sigma1 receptors as compared with [11C]SA4503. In rat brain, similar regional distribution patterns of (+)-[11C]PMV and [11C]SA4503 were shown by tissue dissection and by ex vivo autoradiography. Blocking experiments using (+/-)-PMV, (-)-vesamicol, SA4503, haloperidol and (+/-)-pentazocine showed that the two tracers specifically bound to sigma1 receptors, and that [11C]SA4503 exhibited greater specific binding than (+)-[11C]PMV. No sign of VAChT-specific binding by [11C]SA4503 was observed in the striatum, which is rich in VAChT sites. In conclusion, (+)-[11C]PMV specifically bound to sigma1 receptors in the brain, but to a lesser extent than [11C]SA4503, suggesting that (+)-[11C]PMV is a less preferable PET ligand than [11C]SA4503. On the other hand, the moderate affinity of [11C]SA4503 for VAChT is negligible in vivo.

A simple and rapid radiochemical choline acetyltransferase (ChAT) assay screening test.

A simple radiochemical choline acetyltransferase (ChAT) assay screening test was developed by measuring for [(3)H]acetylcholine ([(3)H]ACh) formed from 0.2 mM [(3)H]acetyl-coenzyme A ([(3)H]acetyl-CoA) and 1 mM choline by 0.2 mg of rat brain homogenates containing ChAT into 96-well microplates. A simple and rapid procedure for isolating [(3)H]ACh from the incubation mixture into 96-well microplates was achieved by using a sodium tetraphenylboron (Kalibor) solution (in ethyl acetate, 0.75%, w/v) and a hydrophobic liquid scintillator mixture (1:5, v/v, 0.2 mL) as an extraction solvent. The benefits of this radiochemical method using 96-well microplates are as follows: (1) this method is reliable and reproducible; (2) many samples can be examined at the same time by this method; (3) this method is economical and effective in reducing radioactive waste. The development of a new simple radiochemical ChAT assay screening test is the first stage of development of radiolabeled ChAT mapping agent.

Synthesis and binding affinities of methylvesamicol analogs for the acetylcholine transporter and sigma receptor.

We synthesized methylvesamicol analogs 13-16 and investigated the binding characteristics of 2-[4-phenylpiperidino]cyclohexanol (vesamicol) and methylvesamicol analogs 13-16, with a methyl group introduced into the 4-phenylpiperidine moiety, to sigma receptors (sigma-1, sigma-2) and to vesicular acetylcholine transporters (VAChT) in membranes of the rat brain and liver. In competitive inhibition studies, (-)-o-methylvesamicol [(-)-OMV] (13) (Ki=6.7 nM), as well as (-)-vesamicol (Ki=4.4 nM), had a high affinity for VAChT. (+)-p-Methylvesamicol [(+)-PMV] (16) (Ki=3.0 nM), as well as SA4503 (Ki=4.4 nM), reported as a sigma-1 mapping agent for positron emission tomography (PET), had a high affinity for the sigma-1 receptor. The binding affinity of (+)-PMV (16) for the sigma-1 receptor (Ki=3.0 nM) was about 13 times higher than that for the sigma-2 (sigma-2) receptor (Ki=40.7 nM). (+)-PMV (16) (Ki=199 nM) had a much lower affinity for VAChT than SA4503 (Ki=50.2 nM) and haloperidol (Ki=41.4 nM). These results showed that the binding characteristics of (-)-OMV (13) to VAChT were similar to those of (-)-vesamicol and that (+)-PMV (16) bound to the sigma-1 receptor with high affinity. In conclusion, (-)-OMV (13) and (+)-PMV (16), which had a suitable structure, with a methyl group for labeling with 11C, may become not only a new VAChT ligand and a new type of sigma receptor ligand, respectively, but may also become a new target compound of VAChT and the sigma-1 receptor radioligand for PET, respectively.

Alternations of static cerebral and systemic circulation in normal humans during 14-day head-down bed rest.

BACKGROUND: There have so far been few reports on the static regulations of cerebral and systemic circulation during prolonged head-down bed rest (HDBR). Our aim was to investigate the time course changes in static cerebral and systemic circulation during 14 days of 6 degrees HDBR. MATERIAL/METHODS: Sixteen subjects participated in the HDBR study. The systolic, mean, and diastolic cerebral blood flow velocities (CBFVs) of the middle cerebral artery were measured using a transcranial Doppler technique. Cerebrovascular bed resistance indices, i.e., resistance index (RI), pulsatility index (PI), and estimated regional cerebrovascular resistance (CVRest) were calculated. The systemic cardiovascular functions, i.e, heart rate (HR), mean arterial pressure (MAP), left ventricular ejection time (LVET), stroke volume (SV), cardiac output (CO), and total peripheral resistance (TPR) were measured or calculated. RESULTS: All CBFVs consistently showed significant decreases from the 2nd day to the last day of the HDBR. The RI and PI showed a rising tendency throughout the HDBR. The CVRest showed significantly higher levels in the later half of the HDBR. The HR and MAP did not change during the HDBR. CONCLUSIONS: The adaptive process of cerebral circulation triggered by HDBR begins very early and leads to a new equilibrium within few days after the onset of HDBR. The alteration of static cerebral circulation with prolonged HDBR, i.e., lowered CBFVs and somewhat higher cerebrovascular bed resistance implies a reduction in the cerebral circulation, but it does not necessarily imply the impaired regulation of cerebral circulation.

Rapid scanning protocol for brain (18)F-FDG PET: a validation study.

UNLABELLED: Although (18)F-FDG PET is an established technique to assess brain glucose use, a shorter imaging time is preferable for patient convenience and increased throughput. The aim of this study was to validate a brain (18)F-FDG PET protocol more rapid than the conventional protocol. METHODS: For comparison of normalized metabolic activities, brain (18)F-FDG PET was performed on 60 healthy subjects and 25 patients with probable Alzheimer's disease (AD), and an additional 20 healthy subjects served as a control group to assess diagnostic performance between the conventional and rapid scanning protocols. Conventional scans were acquired for a total of 20 min (a 10-min emission and a 10-min transmission). Immediately after conventional scanning, rapid scanning was performed for a total of 4 min (a 3-min emission and a 1-min transmission). PET images were anatomically standardized using NEUROSTAT, with pixel values normalized to the individual global value. Two database sets, from the 2 protocols, were compared by regional values and pixel-by-pixel analysis. A receiver-operating-characteristic analysis was performed for comparison of diagnostic accuracy between the 2 protocols. A kinetic simulation study was also performed to examine the possible difference due to the time lag between the protocols. RESULTS: Although small differences in normalized activity were found in several regions in the healthy subjects between the 2 protocols, no significant difference was found in any region in the patient group. The coefficients of variation of the normalized activity were 20%-30% larger in the rapidly scanned images, but the mean z images and their coefficient-of-variation images did not differ. The kinetic simulation study suggested that the differences were caused by the time lag between the 2 protocols. No significant differences were found in area under the receiver-operating-characteristic curves, and the diagnostic accuracies for the detection of AD were virtually equal between the 2 protocols. CONCLUSION: The rapid scanning protocol used in the present study could provide results nearly equivalent to data from the conventional protocol. Thus, it is feasible to use this rapid protocol to detect AD, without losing diagnostic accuracy.

Hypergravity exercise against bed rest induced changes in cardiac autonomic control.

An intermittent exposure to artificial hypergravity with physical exercise by a human centrifuge may provide a countermeasure against various physiological problems after space flight. To test the effects of hypergravity with ergometric exercise on dynamic regulation of heart rate during weightlessness, we quantified autonomic cardiovascular control before and after head-down-tilt bed rest (HDBR) with and without the countermeasure. Twelve male subjects underwent a 14-day period of HDBR. Six of them were exposed to a hypergravity (+1.2 Gz acceleration at heart level) for 30 min with ergometric exercise (60 W, n=4; 40 W, n=2) as a countermeasure on day 1, 2, 3, 5, 7, 9, 11, 12, 13 and 14, during HDBR (CM group). The remaining six were not exposed to a hypergravity exercise during HDBR (control group). Blood pressure and ECG were recorded at a supine position before and after HDBR. The high frequency power of R-R interval (HFRR; 1,008+/-238 to 353+/-56 ms(2) P<0.05) as an index of cardiac parasympathetic activity, and transfer function gain between BP and R-R interval in the high frequency range (GainHF; 21.9+/-5.4 to 14.5+/-4.2 ms/mmHg, P<0.01) as an index of vagally mediated arterial-cardiac baroreflex, decreased significantly after HDBR in the control group. However, these changes were not statistically significant in the CM group (HFRR, 1,150+/-344 to 768+/-385 ms(2); GainHF, 21.5+/-3.3 to 18.6+/-3.4 ms/mmHg). Moreover, baroreflex gain by sequence analysis showed similar results. This observation suggests that the intermittent exposure to hypergravity with ergometric exercise may attenuate the decreases in the parasympathetic activity and the spontaneous arterial-cardiac baroreflex function after weightlessness.

Brain FDG PET study of normal aging in Japanese: effect of atrophy correction.

PURPOSE: The aim of this study was to investigate the effects of atrophy correction on the results of 18F-fluorodeoxyglucose positron emission tomography (FDG PET) in the context of normal aging. METHODS: Before the human study was performed, a Hoffman 3D brain phantom experiment was carried out in order to validate a newly developed correction method for partial volume effects (PVEs). Brain FDG PET was then performed in 139 healthy Japanese volunteers (71 men, 68 women; age 24-81 years). PET images were corrected for PVEs using grey matter volume, which was segmented from co-registered magnetic resonance images and convoluted with the spatial resolution of the PET scanner. We investigated the correlation between advancing age and relative regional FDG activity, which was normalised to the global activity before and after PVE correction using Statistical Parametric Mapping 99. RESULTS: The PET image, when corrected for PVEs, provided more homogeneous tracer distribution in the whole phantom than in the original PET image. The human PET study of both sexes revealed significant negative correlations between age and relative FDG activity in the bilateral perisylvian and medial frontal areas before PVE correction. However, these negative correlations were largely resolved after PVE correction. CONCLUSION: Correction for PVEs was effective in our FDG PET study. The reduction in FDG uptake with advancing age that was detected by FDG PET without PVE correction could be accounted for largely by an age-related cerebral volume loss in the bilateral perisylvian and medial frontal areas.

Functional changes in autonomic nervous system and baroreceptor reflex induced by 14 days of 6 degrees head-down bed rest.

We studied the effects of 14 days of 6 degrees head-down bed rest (HDBR) in 16 healthy male subjects to examine the functional changes in the autonomic nervous system and cardiac baroreceptor reflex response with an emphasis on dynamic changes during HDBR. Beat-by-beat RR intervals (RRIs) and systolic arterial pressures (SAPs) were measured non-invasively from simultaneous, continuous recordings of ECG and arterial pressure waves in supine resting postures. A power spectrum analysis by the fast Fourier transform was applied to a data set composed of interpolated 512 RRIs and 512 SAPs (256 s in duration). Three indices of cardiac baroreceptor reflex sensitivity (BRS) were obtained by applying a sequence technique and a cross-spectrum analysis technique to the spontaneous RRIs and SAPs. The high-frequency band power of RRI variability (HF(RRI)) decreased significantly in the latter part of HDBR and persisted until the initial stage of the post-HDBR period (POST). The low-frequency band power of SAP variability decreased significantly only during the mid-part of HDBR. The BRS(sequence) obtained by the sequence technique showed a significant increase temporarily on the initial day of HDBR. The BRS(sequence) and the estimate of BRS obtained by the cross-spectrum analysis handling the high-frequency band were both significantly decreased on the initial day of POST. Each of the BRS estimates correlated negatively with heart rate and positively with HF(RRI) during HDBR and POST. These results suggest the following: (1) cardiac spontaneous baroreceptor reflex sensitivity might be transiently increased at the initial stage of HDBR, (2) the reduction in vagal modulation on the sinus node occurs from the latter part of HDBR to the initial stage of POST, (3) sympathetic vasomotor control is probably slightly inhibited during the mid-part of HDBR, and (4) the enhancement in cardiac sympathetic modulation and the impairment in cardiac spontaneous baroreceptor reflex sensitivity may occur in the initial stage of POST.

Autonomic cardiovascular changes during and after 14 days of head-down bed rest.

A 14-day, 6 degrees head-down bed rest (HDBR) study was conducted with 12 healthy young men to determine whether there are transient responses of the cardiovascular autonomic regulatory system including cardiovascular, autonomic nervous, and cardiac baroreceptor reflex functions in the acute phases of HDBR and post-HDBR. Compared with the supine position before bed rest, the high-frequency band power (HF(RRI)) of RR intervals (RRIs) decreased significantly at 3, 6, and 24 h of HDBR. This tendency went on until 24 h post-HDBR. Three kinds of cardiac baroreceptor reflex sensitivity (BRS) were estimated from closed-loop approaches to simultaneously recorded spontaneous RRI and systolic arterial pressure (SAP) fluctuations. BRSsequence is based on the simultaneous changes between RRI and SAP. alphaLF and alphaHF are based on a cross-spectrum analysis for low- and high-frequency bands of RRI and SAP. Although BRSsequence decreased significantly at acute phases of both HDBR and post-HDBR, neither alphaLF nor alphaHF decreased significantly at any of the acute phases of HDBR and post-HDBR. Our results suggest that HF(RRI) and BRSsequence can be used effectively to reveal reductions in cardiac vagal nervous modulation on the sinus node and cardiac BRS within 24 h of both HDBR and post-HDBR.

Use of the "supercharge" technique in esophageal and pharyngeal reconstruction to augment microvascular blood flow.

BACKGROUND: Although gastric tubes are commonly used in thoracic esophageal reconstruction, sometimes circumstances are such that an alternative method must be found. On these occasions, pedicled jejunum transfer and colonic interposition are used. After reconstruction, partial necrosis can occur because of poor blood flow in the oral end of the segment used to restore esophageal continuity. To improve this blood flow, we created a technique-the "supercharge" technique-in which we perform additional microvascular blood flow augmentation. METHODS: The supercharge technique was performed in 82 esophageal reconstructions with microvascular blood flow augmentation. Reconstructive methods included a gastric tube in 5 patients, a gastric tube combined with a free jejunal graft in 2, an elongated gastric tube in 6, a pedicled colonic interposition in 26, and a pedicled jejunum in 43. Recipient vessels were located in the neck or chest regions. RESULTS: After microvascular blood flow augmentation, the color and blood flow of the transferred intestine appeared greatly improved. During the operation, thrombosis was noticed in 3 patients and successfully salvaged by reanastomosis. Partial graft necrosis of the oral end of the segment occurred in only 2 patients; 2 patients had anastomotic leakage. CONCLUSIONS: The risk of leakage and partial necrosis of the transferred gut conduit appear to be reduced by using the supercharge technique to augment microvascular blood flow. This reliable technique contributes to the successful reconstruction of esophageal defects.

Changes in bone turnover markers during 14-day 6 degrees head-down bed rest.

Osteoporosis caused by exposure to microgravity represents a serious clinical concern, but the mechanisms have yet to be fully elucidated. The present research aimed to elucidate the effects of microgravity environments on bone turnover, with a specific focus on changes in bone resorption markers such as type I collagen cross-linked N-telopeptides (NTx) and deoxypyridinoline (Dpyr), for which scant data are available regarding detailed time course. Methods using 6 degrees head-down bed rest were utilized to simulate a microgravity environment. Eleven adult male volunteers underwent 6 degrees head-down bed rest for 14 days; measurements were made of serum and urine Ca concentrations, in addition to osteocalcin (OC), bone alkaline phosphatase (ALP), NTx, and Dpyr as bone turnover markers. By the end of bed rest, concentrations of bone ALP had significantly increased, but OC displayed a tendency toward decrease. Concentrations of Dpyr significantly increased from day 6, remaining elevated until the end of bed rest. Concentrations of NTx significantly increased on day 13 and at the end of bed rest. Serum and urinary concentrations of Ca increased significantly at the end of bed rest. Bone ALP represents a relatively early marker of osteoblast differentiation at the matrix maturation phase and OC is a late marker in osteoblast differentiation at the calcification phase. The present results therefore suggest an absolute increase in bone resorption and normal or reduced bone formation, together causing prominent uncoupling and rapid bone loss after simulated microgravity. Moreover, the present results suggest that bone resorption is enhanced at an early stage of exposure to microgravity environments.

[Dynamic cerebral blood flow autoregulation during sevoflurane anesthesia and TIVA].

BACKGROUND: Dynamic cerebral blood flow autoregulation during sevoflurane anesthesia and total intravenous anesthesia (TIVA) is unclear. We examined the cerebral circulation autoregulation during anesthesia by sevoflurane or TIVA. METHODS: We measured mean blood pressure (MBP) and blood flow velocity of the middle cerebral artery by a transcranial Doppler ultrasonography before and during anesthesia using sevoflurane (volatile induction and maintenance of anesthesia (VIMA) group) and using propofol and fentanyl (TIVA group), and the relationship between changes in MBP and cerebral blood flow velocity was evaluated using the method of transfer function analysis. We calculated transfer gain and coherence by cross-spectrum from autospectra of MBP and cerebral blood flow velocity. RESULTS: Transfer gain during anesthesia by TIVA in the low frequency range and high frequency range was near 1 cm.sec-1.mmHg-1. It was about equal to the value of transfer gain before anesthesia. But transfer gain during anesthesia by VIMA was above 2 cm.sec-1.mmHg-1. CONCLUSION: These results suggest that TIVA by propofol and fentanyl maintains the dynamic autoregulation of cerebral blood flow, but sevoflurane impairs the autoregulation.

Follow-up study of upper limb lymphedema patients treated by microsurgical lymphaticovenous implantation (MLVI) combined with compression therapy.

We present a follow-up study of 18 patients with upper limb lymphedema treated by microsurgical lymphaticovenous implantation (MLVI) combined with compression therapy. This combined technique provides increased lymphatic flow through newly created lymphaticovenous bypasses by the MLVI surgery, with assistance for pumping function in the lymphatics by compression therapy. Preoperative assessment of the affected limb was performed by the average enlargement of edema circumference (AEEC), comparing the lymphedema limb and normal limb circumferences. Objective improvement was analyzed by the percent reduction of edema circumference (%REC) at two levels of the lymphedema limb. With an average follow-up of 24 months, 77.8% of patients presented excellent or good results, with %REC >50% at either the distal or proximal site of the treated limb. This combined treatment can be expected to provide favorable long-term results, even for patients with AEEC >8 cm.

Measurement of brain concentration of FK960 for development of a novel antidementia drug: a PET study in conscious rhesus monkeys.

UNLABELLED: This study used PET to measure the time course of the brain concentration of (18)F-labeled N-(4-acetyl-1-piperazinyl)-p-fluorobenzamide monohydrate (FK960), a novel antidementia drug, after oral administration to conscious rhesus monkeys. METHODS: Three young-adult male rhesus monkeys were tested. FK960 (0.1 mg/kg) containing about 370 MBq of (18)F-FK960 was administered orally to each monkey. Dynamic PET images were acquired for 4 h from 5 min after the administration. Arterial blood samples were withdrawn during PET scanning and were analyzed by an automatic well gamma-counter and thin-layer chromatography to determine the time course of authentic (18)F-FK960 activity concentration in plasma. FK960 concentrations in brain and plasma were calculated in units of mol/L using the specific activity of FK960 preparations. RESULTS: (18)F-FK960 penetrated the blood-brain barrier and underwent perfusion-dependent distribution in the entire brain. Maximal concentrations in the brain and plasma were 1.11 +/- 0.30 x 10(-7) mol/L (at 3.0 +/- 0.6 h after administration) and 4.04 +/- 1.29 x 10(-7) mol/L (at 2.0 +/- 1.1 h after administration), respectively. CONCLUSION: We succeeded in measuring the FK960 concentration in the brains of conscious monkeys and in plasma after oral administration at a dose of 0.1 mg/kg. The results suggested that this method can measure the FK960 concentration in the human brain, and a potential use of the PET technique in drug development was demonstrated.

An effective tool for homogeneous assays using a simple robot arm with Stackers-TRISTAN.

The Twister & RapidPlate Integrated System by TANabe, or TRISTAN, consists of a 96-channel dispenser (RapidPlate 96), a plate reader (V-MAX), and a simple robot arm (Twister). We developed TRISTAN for effectively conducting a homogeneous assay. Although this system accommodates fewer than 20 microplates, it has several advantages over conventional robotic systems for high-throughput screening in the following aspects: parameter setting, running time, hardware errors, manpower, and cost-effectiveness. The system proved to be effective and efficient for homogeneous assays.

Parasympathetic nervous activity after administration of atropine and neostigmine using heart rate spectral analysis.

Recently, heart rate spectral analysis has become recognized as a powerful tool for quantitatively evaluating autonomic nervous system activity. The purpose of this study was to analyze parasympathetic nervous activity by heart rate spectral analysis after administration of atropine and neostigmine for reversal of residual neuromuscular blockade. For our study, 36 female patients (26-37 years of age), ASA physical status (PS) I, who were scheduled for laparoscopic examination, were randomly allocated to one of the following four groups: In group A (1∶1), 9 patients received 1.0mg atropine followed 4 min later by 1.0 mg neostigmine. In group B (1∶2), 9 patients received 0.5 mg atropine followed 4 min later by 1.0 mg neostigmine. In group C (1∶2.5), 9 patients received 1.0 mg atropine followed 4 min later by 2.5 mg neostigmine. In group D (1∶2 mix), 9 patients received a mixed solution of atropine 0.5 mg and neostigmine 1.0mg. After finishing the laparoscopic examination, additional anesthesia was maintained with 70% nitrous oxide, 30% oxygen, and 0.5% isoflurane. The control data were obtained 10 min after finishing the laparoscopic examination. After that, the data on atropine were obtained between 2 and 4min after administration of atropine, and the data on neostigmine were obtained between 5 and 7 min after administration of neostigmine. We selected power spectral density of the high-frequency component (HF-p) in heart rate spectral analysis as an index to assess parasympathetic activity. In groups A, B, and C, the HF-p decreased after administration of atropine. In groups B and C, the HF-p increased after administration of neostigmine as compared to the control. In group A, the HF-p increased after neostigmine but did not differ from the control. The difference between groups D and B was not statistically significant. From the results of this study, we concluded that the muscarinic effect of neostigmine could not be sufficiently blocked by atropine at 1/2 dosages of neostigmine, but could be sufficiently blocked by atropine at equivalent dosages of neostigmine, under light isoflurane anesthesia.