The Real-world Efficacy and Safety of Ombitasvir/Paritaprevir/Ritonavir for Hepatitis C Genotype 1.

Objective There are few reports on the outcomes of 12-week paritaprevir, ombitasvir, and ritonavir (PTV/OBV/r) treatment in real-world clinical settings. We aimed to evaluate the efficacy and safety of 12-week treatment with ritonavir-boosted paritaprevir and ombitasvir in patients with hepatitis C virus (HCV) genotype 1 infection in a real-world setting. Methods Fifty-eight patients with chronic hepatitis or compensated hepatic cirrhosis and genotype-1 HCV infection were treated with PTV/OBV/r and followed for 24 weeks after the completion of treatment in 10 centers in northern Tohoku. The efficacy and safety of this 12-week treatment regimen was analyzed. Results Among the 58 treated patients, 18 (31%) had compensated liver cirrhosis, while 11 (19%) patients had experienced treatment failure with another treatment regimen. NS5A resistance-associated variants (RAVs) were detected at baseline in 3 patients (5.2%), including Y93H in two patients and L31M in two patients. One patient had NS5A RAVs at both positions 93 and 31. The overall sustained virological response (SVR) 24 rate was 96.6%. Three patients with NS5A RAVs also achieved an SVR24. The SVR24 rate was not significantly affected by age, sex, prior treatment, prior history of HCC, or liver stiffness. The mean alanine aminotransferase (ALT) levels decreased significantly during this treatment. Adverse events occurred in 15 patients (26%), 26% of which were grade 1 or 2. No severe adverse events occurred. Conclusion In this real-world study, 12-week PTV/OBV/r treatment was effective and safe for treating patients with HCV-1 infection who had chronic hepatitis or compensated hepatic cirrhosis.

Development of a Smartphone App for Visualizing Heart Sounds and Murmurs.

BACKGROUND: Auscultation is one of the basic techniques for the diagnosis of heart disease. However, the interpretation of heart sounds and murmurs is a highly subjective and difficult skill. OBJECTIVES: To assist the auscultation skill at the bedside, a handy phonocardiogram was developed using a smartphone (Samsung Galaxy J, Android OS 4.4.2) and an external microphone attached to a stethoscope. METHODS AND RESULTS: The Android app used Java classes, "AudioRecord," "AudioTrack," and "View," that recorded sounds, replayed sounds, and plotted sound waves, respectively. Sound waves were visualized in real-time, simultaneously replayed on the smartphone, and saved to WAV files. To confirm the availability of the app, 26 kinds of heart sounds and murmurs sounded on a human patient simulator were recorded using three different methods: a bell-type stethoscope, a diaphragm-type stethoscope, and a direct external microphone without a stethoscope. The recorded waveforms were subjectively confirmed and were found to be similar to the reference waveforms. CONCLUSIONS: The real-time visualization of the sound waves on the smartphone may help novices to readily recognize and learn to distinguish the various heart sounds and murmurs in real-time.

Signaling of Prostaglandin E Receptors, EP3 and EP4 Facilitates Wound Healing and Lymphangiogenesis with Enhanced Recruitment of M2 Macrophages in Mice.

Lymphangiogenesis plays an important role in homeostasis, metabolism, and immunity, and also occurs during wound-healing. Here, we examined the roles of prostaglandin E2 (PGE2) receptor (EP) signaling in enhancement of lymphangiogenesis in wound healing processes. The hole-punch was made in the ears of male C57BL/6 mice using a metal ear punch. Healing process and lymphangiogenesis together with macrophage recruitment were analyzed in EP knockout mice. Lymphangiogenesis was up-regulated in the granulation tissues at the margins of punched-hole wounds in mouse ears, and this increase was accompanied by increased expression levels of COX-2 and microsomal prostaglandin E synthase-1. Administration of celecoxib, a COX-2 inhibitor, suppressed lymphangiogenesis in the granulation tissues and reduced the induction of the pro-lymphangiogenic factors, vascular endothelial growth factor (VEGF) -C and VEGF-D. Topical applications of selective EP receptor agonists enhanced the expressions of lymphatic vessel endothelial hyaluronan receptor-1 and VEGF receptor-3. The wound-healing processes and recruitment of CD11b-positive macrophages, which produced VEGF-C and VEGF-D, were suppressed under COX-2 inhibition. Mice lacking either EP3 or EP4 exhibited reduced wound-healing, lymphangiogenesis and recruitment of M2 macrophages, compared with wild type mice. Proliferation of cultured human lymphatic endothelial cells was not detected under PGE2 stimulation. Lymphangiogenesis and recruitment of M2 macrophages that produced VEGF-C/D were suppressed in mice treated with a COX-2 inhibitor or lacking either EP3 or EP4 during wound healing. COX-2 and EP3/EP4 signaling may be novel targets to control lymphangiogenesis in vivo.

Trial of real-time locating and messaging system with Bluetooth low energy.

BACKGROUND: Hospital real-time location systems (RTLS) are increasing efficiency and reducing operational costs, but room access tags are necessary. OBJECTVE: We developed three iPhone 5 applications for an RTLS and communications using Bluetooth low energy (BLE). METHODS: The applications were: Peripheral device tags, Central beacons, and a Monitor. A Peripheral communicated with a Central using BLE. The Central communicated with a Monitor using sockets on TCP/IP (Transmission Control Protocol/Internet Protocol) via a WLAN (wireless local area network). To determine a BLE threshold level for the received signal strength indicator (RSSI), relationships between signal strength and distance were measured in our laboratory and on the terrace. RESULTS: The BLE RSSI threshold was set at -70 dB, about 10 m. While an individual with a Peripheral moved around in a concrete building, the Peripheral was captured in a few 10-sec units at about 10 m from a Central. The Central and Monitor showed and saved the approach events, location, and Peripheral's nickname sequentially in real time. Remote Centrals also interactively communicate with Peripherals by intermediating through Monitors that found the nickname in the event database. CONCLUSIONS: Trial applications using BLE on iPhones worked well for patient tracking, and messaging in indoor environments.

Effects of sodium bisulfite with or without procaine derivatives on axons of cultured mouse dorsal root ganglion neurons.

BACKGROUND AND OBJECTIVES: Sodium bisulfite (NaHSO3) was clinically used as a preservative agent for local anesthetics but was later suspected to be neurotoxic. However, recent studies reported that NaHSO3 reduces the neurotoxicity of local anesthetics. The purpose of this study was to examine the effects of NaHSO3 with and without procaine on axonal transport in cultured mouse dorsal root ganglion (DRG) neurons. METHODS: Experiment 1 served to determine the dose-dependent effects of NaHSO3 on axonal transport (DRG neurons were treated with 0.01, 0.1, 1, 10, or 20 mM of NaHSO3), whereas experiment 2 investigated the effect of 0.1 mM NaHSO3 on the action of local anesthetics on axonal transport (DRG neurons were treated with 1 mM procaine alone, or with 0.1 mM NaHSO3 plus 1 mM procaine). As an additional experiment, DRG neurons were also treated with 1 mM chloroprocaine alone, or with 0.1 mM NaHSO3 plus 1 mM chloroprocaine. In these experiments, we analyzed the percent change in the number of anterogradely and retrogradely transported organelles and recorded changes in neurite morphology using video-enhanced microscopy. RESULTS: In experiment 1, NaHSO3 at more than 1 mM caused cell membrane damage and complete inhibition of axonal transport, whereas 0.1 mM NaHSO3 maintained axonal transport at 40% to 60% of control with intact cell membrane. In experiment 2, 1 mM procaine alone maintained axonal transport at 90% to 100%. However, application of 1 mM procaine-0.1 mM NaHSO3 solution resulted in deformation of neurites and with complete cessation of axonal transport. Likewise, although 1 mM chloroprocaine maintain axonal transport at 80% to 100%, 1 mM chloroprocaine-0.1 mM NaHSO3 arrested axonal transport. CONCLUSIONS: NaHSO3 resulted in a dose-dependent damage to the cell membrane and axonal transport, especially when used in combination with procaine or chloroprocaine.

Telomere lengths at birth in trisomies 18 and 21 measured by Q-FISH.

Trisomies 18 and 21 are genetic disorders in which cells possess an extra copy of each of the relevant chromosomes. Individuals with these disorders who survive birth generally have a shortened life expectancy. As telomeres are known to play an important role in the maintenance of genomic integrity by protecting the chromosomal ends, we conducted a study to determine whether there are differences in telomere length at birth between individuals with trisomy and diploidy, and between trisomic chromosomes and normal chromosomes. We examined samples of peripheral blood lymphocytes (PBLs) from 31 live neonates (diploidy: 10, trisomy 18: 10, trisomy 21: 11) and estimated the telomere length of each chromosome arm using Q-FISH. We observed that the telomeres of trisomic chromosomes were neither shorter nor longer than the mean telomere length of chromosomes as a whole among subjects with trisomies 18 and 21 (intra-cell comparison), and we were unable to conclude that there were differences in telomere length between 18 trisomy and diploid subjects, or between 21 trisomy and diploid subjects (inter-individual comparison). Although it has been reported that telomeres are shorter in older individuals with trisomy 21 and show accelerated telomere shortening with age, our data suggest that patients with trisomies 18 and 21 may have comparably sized telomeres. Therefore, it would be advisable for them to avoid lifestyle habits and characteristics such as obesity, cigarette smoking, chronic stress, and alcohol intake, which lead to marked telomere shortening.

Pancreatic agenesis with congenital diaphragmatic hernia and congenital heart disease: a case report.

Pancreatic agenesis is a rare disease that causes neonatal diabetes mellitus and exocrine pancreatic insufficiency. We report the case of a very low birth weight infant with congenital diaphragmatic hernia, and congenital heart disease (ventricular septal defect and patent ductus arteriosus). Failure to gain weight, despite well-managed respiratory and heart failure, was improved by infusion of subcutaneous insulin, administration of pancreatic enzyme, and nutrition of medium-chain-triglyceride -enriched formula. This is the first case of pancreatic agenesis with both malformations where the patient is discharged from the hospital. Early diagnosis and adequate treatments to compensate pancreatic function may prevent mortality and improve growth.

Neurotropin inhibits axonal transport in cultured mouse dorsal root ganglion neurons.

Axonal transport is a basic neuronal cell function and important for the supply of materials that maintain neuronal cells, and any increase or decrease in axonal transport expresses the state of neurons. Neurotropin is an analgesic agent commonly used for the treatment of chronic pain, but its mechanism of action remains not fully understood. The effects of neurotropin have been investigated in various animal models of nerve injury and chronic pain. In the present study, we dissected the effects of neurotropin on sensory neurons with a special focus on axonal transport using cultured mouse dorsal root ganglion (DRG) neurons. Movement of organelles in neurites was recorded by real-time video-enhanced microscopy. Neurotropin significantly reduced bidirectional axonal transport in time- and concentration-dependent manners without affecting the diameter of these neurites. This is the first report to show the inhibitory effect of neurotropin on axonal transport, and suggest that this action may mediate, at least in part, the analgesic effects of this agent.

Dextro-transposition of the great arteries in a neonate with ventricular septal defect and pulmonary stenosis complicated by premature closure of the ductus arteriosus and possible pathophysiology of both defects.

Premature closure of the ductus arteriosus (PCDA) and transposition of the great arteries (TGA) cause persistent pulmonary hypertension of the newborn (PPHN). We present a case of a newborn who demonstrated d-TGA with ventricular septal defect (VSD) and pulmonary stenosis (PS) complicated by PCDA. The neonate showed severe cyanosis resistant to resuscitation soon after birth, and was diagnosed with d-TGA with VSD by echocardiography. PPHN was also suspected based on physical symptoms and results of echocardiography. The neonate was given inhaled nitric oxide, prostaglandin E1, and catecholamines under sedation, and underwent a balloon atrial septostomy (BAS). His condition gradually improved, and he was extubated on day 7, but his pulmonary subvalvular stenosis gradually worsened and pulmonary blood flow was markedly decreased. A second BAS was performed on day 27 and he showed no improvement. Blalock-Taussig shunt surgery was performed on day 34, which markedly improved his condition. The co-existence of d-TGA and PCDA is generally a lethal state. In our patient, an increase in pulmonary blood flow during the fetal period was restricted because of PS and outlet flow from the left ventricle to the right ventricle via the VSD. This restricted blood flow through the ductus arteriosus, which led to narrowing of the DA. At the same time, damage to and constrictive changes of the pulmonary vessels were prevented. The ductus arteriosus should be carefully evaluated to exclude PCDA in cases of d-TGA. The presence of both VSD and PS may be a prognostic factor in such cases.

Effects of indomethacin on patent ductus arteriosus in neonates with genetic disorders and/or congenital anomalies.

OBJECTIVES: The study aimed to evaluate the effectiveness of intravenous indomethacin (IND) therapy for patent ductus arteriosus (PDA) in neonates with genetic disorders and/or congenital anomalies soon after birth. STUDY DESIGN: A total of 301 neonates with a genetic disorder and/or congenital anomalies and with a gestational age of ≥ 35 weeks were admitted during the study period. Eighty-five neonates with 56 genetic disorders (30 cases of trisomy 21, 10 cases of trisomy 18, and 16 others) and 29 congenital anomalies, and with clinical symptoms received intravenous IND therapy. The management methods were similar to those used for PDA in low-birth-weight infants. RESULTS: IND therapy had a clinical benefit at a high rate of 79% in these patients (90% and 70% in neonates with trisomies 21 and 18, respectively), including complete closure of the PDA in 52% of the patients. Although oliguria was observed in 43 infants (51%) and slight gastrointestinal bleeding was observed in 12 (14%), no infants had severe complications such as intracranial bleeding. CONCLUSIONS: IND therapy is an effective treatment option before considering surgery for PDA in neonates with genetic disorders and/or congenital anomalies. This therapy may reduce the difficulty of treatment in the acute stage among these neonates.

Examination of the percentage of immature platelet fraction in term and preterm infants at birth.

BACKGROUND: Reticulated platelets (RPs) are newly synthesized platelets. Recently, an automatic method was established to detect RPs as a percentage of the immature platelet fraction (IPF%). Although, neonates often develop thrombocytopenia at some time during their hospitalization, the details of IPF% in neonates remain unclear. We, therefore, studied the relations between IPF% and other factors to gain a more detailed understanding of IPF% in neonates. METHODS: The following clinical data were obtained from the medical records of 105 neonates who met our inclusion criteria: Gestational age, birth weight, IPF% and platelet count of neonatal peripheral blood at birth, and perinatal data. The subjects were divided into three groups: Group A, birth weight standard deviation score (SDS) ≥ -2 standard deviation (SD) and ≤ +2 SD; Group S, < -2 SD; and Group L, > +2 SD. RESULTS: IPF% correlated negatively with platelet count at birth in the whole study population. IPF% was 2.8 ± 1.3% in term neonates, and IPF correlated negatively with gestational age and birth weight. Platelet count correlated positively with birth weight SDS in the whole study population and in Group S. IPF% correlated negatively with birth weight SDS in the whole study population and in Group S. In neonates with a platelet count below 25 × 10(4)/µl, IPF% correlated negatively with platelet count. Among other neonates, however, IPF% remained almost constant. CONCLUSION: Monitoring of IPF% is useful for estimating the function of thrombocytopoiesis in neonates and preterm infants.

Effect of inhibition of superoxide dismutase on motor neurons during growth: comparison of phosphorylated and non-phosphorylated neurofilament-containing spinal neurons by histogram distribution.

We reported recently that non-phosphorylated neurofilaments (NF)-positive neurons were more sensitive to the growth inhibitory effects of Cu/Zn superoxide dismutase (SOD1) than phosphorylated NF-positive neurons. The findings suggested that non-phosphorylated NF-positive neurons, presumed to represent spinal motor neurons, are more vulnerable to oxidative stress than other neurons, and thus explain in part the selective degeneration of motor neurons in amyotrophic lateral sclerosis. The present investigation is an extension to our previous study and examined the neurite growth process in the presence of diethyldithiocarbamate (DDC), an SOD1 inhibitor. Non-phosphorylated NF, representing spinal motor neurons, and phosphorylated NF, representing other spinal neurons, were stained with SMI-32 and SMI-31 antibodies, respectively. The distribution histogram of neurite length after treatment with 0 nM DDC (control) for 72 h appeared flatter compared with that of 24h. Although the addition of DDC (1 nM, 10 nM, 100 nM, or 1000 nM) to the culture medium for 72 h shifted the histogram of neurite length to a shorter range in a concentration-dependent manner, the neurite of SMI-31-immunoreactive neurons grew under DDC. On the other hand, DDC-treatment for 72 h altered the neurite growth of SMI-32-immunoreactive neurons compared with that for 24-h. The results suggest that SOD1 inhibition, representing accumulation of endogenous oxidative stress, suppresses neurite growth of spinal motor neurons, and that the growth of spinal motor neurons is more sensitive to oxidative stress than other types of neurons.

Immunocytochemical colocalization of fibroblast growth factor-1 with neurotrophin-3 in mouse alveolar macrophages.

Alveolar macrophages are known to express a variety of growth factors and neurotrophins. Fibroblast growth factor-1 (FGF-1) is abundantly present in the lung and has mitogenic and neurotrophic activities similarly to neurotrophins. In order to determine whether FGF-1 associates with neurotrophins in alveolar macrophages, we investigated the immunocytochemical colocalization of FGF-1 with neurotrophins, nerve growth factor (NGF), brain-derived neurotrophic factor (BDNF), and neurotrophin-3 (NT-3), in mouse alveolar macrophages. The results showed that 34% of macrophages were immunoreactive for FGF-1, 10% for NGF, 9% for BDNF, and 17% for NT-3. Of FGF-1-immunoreactive (IR) macrophages, 16% were immunoreactive for NT-3, but only small percentages were immunoreactive for NGF (0.8%) and for BDNF (0.3%). FGF-1 and neurotrophins were all localized in the intracellular vesicles. In the vesicles, FGF-1 and NT-3 were frequently colocalized. All macrophages expressed lysosome-associated protein-2 (LAMP-2), a late endosomal and lysosomal marker, and early endosomes antigen 1 (EEA1), an early endosomal marker. FGF-1 and NT-3 were predominantly colocalized with LAMP-2 rather than with EEA1, whereas NGF and BDNF were colocalized with EEA1 rather than with LAMP-2. These results indicate that FGF-1 and NT-3 are substantially expressed in mouse alveolar macrophages and colocalized in vesicles, predominantly in late endosomes and lysosomes.

Intrathecally administered ropivacaine is less neurotoxic than procaine, bupivacaine, and levobupivacaine in a rat spinal model.

PURPOSE: The aim of this study was to compare the neurotoxicity of intrathecal procaine, bupivacaine, levobupivacaine, and ropivacaine in an animal model. METHODS: The study comprised two experiments. In the concentration experiment, rats (n = 78) were administered 0.12 µL·g(-1) body weight (BW) of 2% or 20% procaine, 0.5% or 5% bupivacaine, 0.5% or 5% levobupivacaine, or 0.5% or 5% ropivacaine. Based on the findings, the doses were increased by volume in the subsequent volume experiment using 0.12, 0.24, or 0.48 µL·g(-1) BW of 6% procaine, 6% levobupivacaine, or 6% ropivacaine (n = 79). Walking behaviour and sensory threshold were analyzed, and a histological examination of the spinal cord, posterior and anterior roots, and cauda equina was performed. RESULTS: The concentration experiment showed abnormalities only in the 5% bupivacaine group, and these abnormal findings were in the posterior root (PR) and posterior column (PC). The volume experiment revealed that procaine 0.24 µL·g(-1) was neurotoxic, mainly affecting the PR. At 0.48 µL·g(-1), severe injury was observed in the PR and PC in all six procaine rats and four of six levobupivacaine rats, while milder injury was limited to the PR in one of six ropivacaine rats, which differed significantly from the former two groups (P = 0.006 and P = 0.014, respectively). Electron microscopy showed axonal degeneration. CONCLUSION: All four local anesthetics seemed to cause identical neurotoxic lesions commencing in the PR and extending to the PC by axonal degeneration. Bupivacaine appeared to be the most neurotoxic of the four drugs, and the neurotoxicity at higher doses increased by volume with procaine > levobupivacaine > ropivacaine.

Long survival of congenital alveolar capillary dysplasia patient with NO inhalation and epoprostenol: effect of sildenafil, beraprost and bosentan.

The case is described herein of a patient with alveolar capillary dysplasia with double-outlet right ventricle and duodenal atresia who survived for a remarkably long time. The newborn girl was born at a gestational age of 36 weeks and weighed 1926 g. One min after delivery the Apgar score was 4. The patient had persistent pulmonary hypertension (PH) and needed nitric oxide inhalation and i.v. epoprostenol all through her life. Although other oral medications for PH were tried, they could not be used in practice because of gastrointestinal complications. The patient died on the 237 th day of life as a result of worsening PH associated with infection.

Inhibition of superoxide dismutase selectively suppresses growth of rat spinal motor neurons: comparison with phosphorylated neurofilament-containing spinal neurons.

Amyotrophic lateral sclerosis (ALS) is characterized by selective degeneration of motor neurons. The reason why only motor neurons are targeted is unknown. Since ALS has been linked to mutations in Cu/Zn superoxide dismutase (SOD1), oxidative stress is regarded as a major cause of ALS. We hypothesized that motor neurons are more susceptible to oxidative stress than other neurons. To test our hypothesis, we investigated differences in neurite growth between motor and non-motor neurons under SOD1 inhibition. Spinal motor neurons were identified by immunocytochemistry using anti-non-phosphorylated neurofilament (NF) antibody (SMI-32). Other neurons immunoreactive to an antibody against phosphorylated NF (SMI-31) were used as a control. Cultured rat spinal neurons were treated with the SOD1 inhibitor diethyldithiocarbamate (DDC). SMI-32-immunoreactive neurons were more sensitive to the growth inhibitory effects of DDC than SMI-31-immunoreactive neurons. Such inhibition was blocked by the antioxidants, L-ascorbic acid, L-histidine, astaxanthin, α-tocopherol, and ß-carotene. The results suggested that spinal motor neurons are more vulnerable to oxidative stress than other neurons, which may explain in part the selective degeneration of motor neurons in ALS.

Characteristics of weak base-induced vacuoles formed around individual acidic organelles.

We have previously found that the weak base 4-aminopyridine induces Brownian motion of acidic organelles around which vacuoles are formed, causing organelle traffic disorder in neurons. Our present study investigated the characteristics of vacuoles induced by weak bases (NH(4)Cl, aminopyridines, and chloroquine) using mouse cells. Individual vacuoles included acidic organelles identified by fluorescent protein expression. Mitochondria and actin filaments were extruded outside the vacuoles, composing the vacuole rim. Staining with amine-reactive fluorescence showed no protein/amino acid content in vacuoles. Thus, serous vacuolar contents are probably partitioned by viscous cytosol, other organelles, and cytoskeletons, but not membrane. The weak base (chloroquine) was immunochemically detected in intravacuolar organelles, but not in vacuoles. Early vacuolization was reversible, but long-term vacuolization caused cell death. The vacuolization and cell death were blocked by the vacuolar H(+)-ATPase inhibitor and Cl--free medium. Staining with LysoTracker or LysoSensor indicated that intravacuolar organelles were strongly acidic and vacuoles were slightly acidic. This suggests that vacuolization is caused by accumulation of weak base and H(+) in acidic organelles, driven by vacuolar H(+)-ATPase associated with Cl(-) entering, and probably by subsequent extrusion of H(+) and water from organelles to the surrounding cytoplasm.

Inhibition of axonal transport caused by tert-butyl hydroperoxide in cultured mouse dorsal root ganglion neurons.

Reactive oxygen species (ROS) are capable of affecting neuronal cell function and structure. Here, we investigated the direct effects of hydrogen peroxide (H(2)O(2)), one of the ROS, on axonal transport in cultured mouse dorsal root ganglion neurons using video-enhanced microscopy. Treatment of neurons with the H(2)O(2) donor tert-butyl hydroperoxide (TBHP; 10 nM-1 mM) inhibited anterograde and retrograde movement of organelles in a time- and concentration-dependent manner. Mitochondria and lysosomes were clearly swollen by TBHP at 100 µM and 1 mM in association with complete and irreversible cessation of axonal transport. In contrast, cytoskeletal structures were apparently unchanged even at the highest TBHP concentration (1 mM). Lipid peroxides, detected by swallow-tailed perylene derivative fluorescence, were produced by TBHP in plasma membranes and more highly in organelle membranes. The TBHP-induced inhibition of axonal transport, lipid peroxide production, and organelle swelling were blocked by pretreatment with α-tocopherol (vitamin E, 1 mM). These results suggest that H(2)O(2) inhibited axonal transport via lipid peroxidation along with degenerative changes in organelles.

Effects of 4-aminopyridine on organelle movement in cultured mouse dorsal root ganglion neurites.

Aminopyridines, widely used as a K(+) channel blocker, are membrane-permeable weak bases and have the ability to form vacuoles in the cytoplasm. The vacuoles originate from acidic organelles such as lysosomes. Here, we investigated the effects of 4-aminopyridine (4-AP) on organelle movement in neurites of cultured mouse dorsal root ganglion (DRG) neurons by using video-enhanced microscopy. Some experiments were carried out using fluorescent dyes for lysosomes and mitochondria and confocal microscopy. Treatment of DRG neurons with 4 mM 4-AP caused Brownian movement of some lysosomes within 5 min. The Brownian movement gradually became rapid and vacuoles were formed around individual lysosomes 10-20 min after the start of treatment. Axonal transport of organelles was inhibited by 4-AP. Lysosomes showing Brownian movement were not transported in longitudinal direction of the neurite and the transport of mitochondria was interrupted by vacuoles. The 4-AP-induced Brownian movement of lysosomes with vacuole formation and inhibition of axonal transport were prevented by the simultaneous treatment with vacuolar H(+) ATPase inhibitor bafilomycin A1 or in Cl(-)-free SO(4)(2-) medium. These results indicate that changes in organelle movement by 4-AP are related to vacuole formation and the vacuolar H(+) ATPase and Cl(-) are required for the effects of 4-AP.

Spinal procaine is less neurotoxic than mepivacaine, prilocaine and bupivacaine in rats.

BACKGROUND AND OBJECTIVES: Lidocaine has been reported to be more neurotoxic than other local anesthetics. Alternatives to lidocaine with lower toxicity and shorter duration of action are desirable. Therefore, we compared the histologic and functional changes induced by intrathecal injection of prilocaine, mepivacaine, procaine, and bupivacaine in rats. METHODS: Rats (n = 184) randomly received via an intrathecal catheter 0.12 microL/g body weight of 2%, 10%, 16%, or 20% prilocaine, mepivacaine, or procaine; 0%, 0.5%, 2.5%, 4%, or 5% bupivacaine in distilled water; or distilled water or 15% glucose solution alone as a control. We evaluated neurofunction by analyzing walking behavior and sensory threshold and examined the L3 spinal cord, posterior and anterior roots, and cauda equina by light and electron microscopy. RESULTS: The recovery time to normal ambulation after intrathecal injection was significantly faster with procaine than with the other 3 drugs at all concentrations. There were no significant differences in the sensory threshold among the 4 anesthetics. Histologic damage was observed only in rats treated with greater than 16% prilocaine or mepivacaine or with greater than 4% bupivacaine. Histologic damage occurred at the posterior root and posterior white matter and was characterized by axonal degeneration. Rats treated with procaine, even at 20%, showed no histologic abnormalities. CONCLUSION: In this animal model, the neurotoxicity of intrathecal procaine was the mildest, and the recovery time to ambulation with procaine was the fastest among the 4 tested anesthetics.