Repeated administration of acrylamide for 28 days suppresses adult neurogenesis of the olfactory bulb in young-adult rats.

Acrylamide (AA) is a neurotoxicant that inhibits synaptic function in distal axons. We previously found that AA decreased neural cell lineages during late-stage differentiation of adult hippocampal neurogenesis and downregulated genes related to neurotrophic factor, neuronal migration, neurite outgrowth, and synapse formation in the hippocampal dentate gyrus in rats. To investigate whether olfactory bulb (OB)-subventricular zone (SVZ) neurogenesis is similarly affected by AA exposure, AA was administered to 7-week-old male rats via oral gavage at doses of 0, 5, 10, and 20 mg/kg for 28 days. Immunohistochemical analysis revealed that AA decreased the numbers of doublecortin-positive (+) cells and polysialic acid-neural cell adhesion molecule+ cells in the OB. On the other hand, the numbers of doublecortin+ cells and polysialic acid-neural cell adhesion molecule+ cells in the SVZ did not change with AA exposure, suggesting that AA impaired neuroblasts migrating in the rostral migratory stream and OB. Gene expression analysis in the OB revealed that AA downregulated Bdnf and Ncam2, which are related to neuronal differentiation and migration. These results suggest that AA decreased neuroblasts in the OB by suppressing neuronal migration. Thus, AA decreased neuronal cell lineages during late-stage differentiation of adult neurogenesis in the OB-SVZ, similar to the effect on adult hippocampal neurogenesis.

Repeated administration of acrylamide for 28 days reduces late-stage progenitor cells and immature granule cells accompanying impaired neurite outgrowth in the adult hippocampal neurogenesis in young-adult rats.

Acrylamide (AA) is a neurotoxicant that causes synaptic impairment in distal axons. We previously found that developmental exposure to AA decreased proliferation of late-stage neural progenitor cells (NPCs) in the hippocampal neurogenesis of the dentate gyrus (DG) in rats. To investigate whether hippocampal neurogenesis is similarly affected by AA exposure in a general toxicity study, AA was administered to 7-week-old male rats via oral gavage at dosages of 0, 5, 10, and 20 mg/kg for 28 days. In the subgranular zone (SGZ) and granule cell layer, AA decreased the densities of doublecortin-positive (+) cells and TOAD-64/Ulip/CRMP protein 4b+ cells per SGZ length. In addition, AA decreased the neurite length of doublecortin+ cells and downregulated genes related to neurite outgrowth (Ncam2 and Nrep) and neurotrophic factor (Bdnf and Ntrk2) in the DG. These results suggest that AA exposure for 28 days decreases type-3 NPCs and immature granule cells in neurogenesis of granule cell lineages involving the impairment of neurite outgrowth in young-adult rats. In the DG hilus, AA increased the density of cholinergic receptor nicotinic beta 2 subunit+ cells. AA also downregulated Reln related to the control of neuronal migration by interneurons in the DG. Furthermore, AA decreased the density of glial fibrillary acidic protein (GFAP)+ astrocytes in the DG hilus and downregulated Gfap and the genes of oligodendrocyte progenitor cells (Cspg4 and Pdgfra). Thus, AA decreased granule cell lineage subpopulations in the late-stage differentiation of hippocampal neurogenesis after young-adult stage exposure, exhibiting a pattern similar to the developmental exposure.

Corneal lesions related to an anesthetic mixture of medetomidine, midazolam, and butorphanol treatment in rats.

An anesthetic mixture of medetomidine, midazolam and butorphanol (MMB) has been recently used in laboratory animals. We observed corneal opacity in nephrectomized rats that had undergone two operations under MMB anesthesia at 4 and 5 weeks of age. To evaluate the features of this corneal opacity, ophthalmic examinations were conducted in 83 nephrectomized rats, and 8 representative animals with corneal opacity were evaluated histopathologically 4 weeks after operation. The ophthalmic examinations revealed that 66/83 animals had corneal opacity, which was characterized histopathologically by mineralization with or without inflammation in the corneal stroma. In addition, to examine the possible causes of this corneal opacity, we investigated whether similar corneal changes were induced by the MMB anesthetic treatment in normal rats. The MMB anesthetic was administered twice to 4- and 5-week-old normal SD rats (5 animals/age) in the same manner as for the nephrectomized rats. Ophthalmic examinations were conducted in all the animals once a week, and the animals were necropsied 4 weeks after the first administration. In normal rats, similar corneal opacity was observed after the first administration, and increases in the severity and size of the corneal opacity were noted after the second administration. In conclusion, this study revealed the features of corneal opacity in rats undergoing nephrectomy under MMB anesthesia and the occurrence of similar corneal opacity in normal rats treated with MMB anesthetic. To the best of our knowledge, this is the first report of corneal opacity related to MMB anesthetic treatment in rats.

Drug-induced lenticular opacity and accumulation of cholesterol-related substances in the lens cortex of dogs.

TP0446131, developed as an antidepressant agent, was found to cause lenticular opacity in a 13-week repeated-dose study in dogs. Histopathologically, the lenticular opacity was observed as a degeneration of the lens fibers, characterized by irregularity in the ordered arrangement of the fibers which is necessary to maintain the transparency of the lens, and was considered to manifest clinically as cataract. To evaluate the development mechanism of the lenticular opacity, the chemical constituents of the lens, which is known to be associated with the development of cataract, were examined. The results of liquid chromatography-tandem mass spectrometry analysis revealed an increase in the amplitudes of 3 unknown peaks in a dose- and time-dependent manner in the lens, with no remarkable changes in the other chemical components tested. In addition, the content of cholesterol, alterations of which have been reported to be associated with cataract, remained unchanged. The mass spectral data and chromatographic behavior of the 3 peaks indicated that these peaks corresponded to sterol-related substances, and that one of them was 7-dehydrocholesterol, a precursor of cholesterol biosynthesis. This finding suggested that TP0446131 exerts some effects on the cholesterol biosynthesis pathway, which could be involved in the development of the cataracts. Furthermore, increases in the levels of these sterol-related substances were also detected in the serum, and were, in fact, noted prior to the onset of the cataract, suggesting the possibility that these substances in the serum could be used as potential safety biomarkers for predicting the onset of cataract induced by TP0446131.

Spontaneous Ocular Abnormalities in Sprague-Dawley Rats.

We collected historical control data derived from pretreatment ophthalmologic examinations of young (4 to 7 wk of age) Sprague-Dawley (Crl:CD[SD]) male, (2033, 42 lots) and female (1322, 32 lots) rats used in toxicity studies at our facility from 2004 through 2015. Ophthalmologic examination of male and female rats by using a binocular indirect ophthalmoscope and slit lamp revealed high incidences of corneal opacity (61% and 60%, respectively), lenticular opacity (43% and 47%), persistent hyaloid artery (21% and 17%), and retinal folds (27% and 27%). All other ocular abnormalities of the globe, conjunctiva, cornea, anterior chamber, lens, iris, vitreous, and choroid or retina occurred at incidences of less than 5%. Corneal opacities were localized mainly in the corneal nasal (38% and 37%) and paracentral (32% and 33%) areas, and lenticular opacities predominantly occurred in the nuclear area (31% and 34%). We then compared the incidences of spontaneous ocular abnormalities between the first (2004 through 2009) and second (2010 through 2015) 6-y periods. Corneal opacity and persistent hyaloid artery in male and female rats occurred more frequently during the second 6-y than during the first (corneal opacity, second period: male, 68%; female, 66%; corneal opacity, first period: 49% and 51%; persistent artery, second period, 26% and 23%; persistent artery, first period; 12% and 10%). These results support the importance of updating historical control data regularly and providing useful information for toxicologists and ophthalmologists to differentiate treatment-related changes from spontaneous lesions.

[ST Segment Elevation on Electrocardiogram (ECG) during Pleural Lavage with Distilled Water;Report of a Case].

Pleural lavage with distilled water is often employed in lung resection to eliminate malignant cells. Here we report a case of transient ST segment elevation on electrocardiogram (ECG) during pleural lavage with distilled water. A 73-year-old female was referred to our hospital because of an abnormal shadow on a chest roentogenogram. Chest computed tomography scan revealed a mass in left S4+5 segment of left upper lobe. It was proved to be adenocarcinoma of the lung by transbronchial lung biopsy and she underwent left upper lobectomy. During pleural lavage with distilled water, ST segment was elevated on ECG. In this case, it was because that the pericardium was excised and the myocardium was exposed to distilled water during pleural lavage.

Gene expression analysis in the lung of the rasH2 transgenic mouse at week 4 prior to induction of malignant tumor formation by urethane and N-methylolacrylamide.

The rasH2 transgenic (Tg) mice are susceptible to genotoxic and some non-genotoxic carcinogens. In carcinogenicity studies carried out using rasH2 Tg mice, the carcinogenic potential of chemicals are evaluated over a 26-week experimental period. In the present study, we examined the comprehensive gene expressions in the lungs of Tg and non-Tg mice prior to the induction of malignant tumors. Urethane (UR), a mutagenic carcinogen, was administered for 4 weeks, and thereafter withdrawn for 22 weeks. N-methylolacrylamide (NMA), a non-mutagenic carcinogen, was administered for 26 weeks. At week 4, gene expression analysis of non-neoplastic part of the lungs demonstrated changes in the expressions of the cell-cycle and inflammation related genes following UR and NMA treatment, respectively, in both the Tg and non-Tg mice. The gene expressions of epireguline, aurora kinase B, and cyclin B1 increased in the UR-treated Tg mice. We also found an increase in the plasma carcinoembryonic antigen level in the UR-treated Tg mice. Although UR treatment induced the formation of adenomas or adenocarcinomas in the lungs in all mice, earlier induction was apparent in the Tg mice. NMA treatment was found to induce the formation of adenomas and adenocarcinomas at week 26 in the Tg mice, but not in the non-Tg mice, and no expressions of specific genes were apparent in either genotype of mice. Our results indicate that analysis of cancer-related gene expressions in the lungs and plasma biomarkers at week 4 in rasH2 Tg mice could be a screening tool for carcinogenicity, especially of mutagenic carcinogens.

[Perioperative Effect of Intraoperative Fluid Restriction with the Same Fixed Volume of 5 ml · kg(-1) · h(-1) in Patients Undergoing Major Abdominal Versus Thoracic Surgery].

BACKGROUND: Major abdominal surgery accompanies the higher magnitude of physiological stress response and may require an additional replacement fluid for the redistributed volume. Intraoperative volume restriction strategy is recommended to avoid fluid overload leading to increased mortality. We conducted a comparative study of the perioperative effects of intraoperative fluid restriction in abdominal versus thoracic surgery. METHODS: Each 15 patients having major abdominal or thoracic surgery were studied prospectively. All participants were identically given intraoperative iv crystalloid of 5 ml · kg(-1) · hr(-1) under combined epidural/general anesthesia. Plasma level of AVP, aldosterone, angiotensin II and IL-6 as well as body water composition by bioelectrical impedance analysis was examined at preoperative period, at the end of surgery and on the first postoperative day. RESULTS: In abdominal surgery group there was significantly less intraoperative urine output compared with thoracic surgery group. No significant differences were found between two groups in extracellular water volume chnages, AVP, aldosterone angiotensin II, IL-6 level and postoperative renal function. CONCLUSIONS: Restrictive fluid therapy with intraoperative crystalloid of 5 ml · kg(-1) · hr(-1) can be safely used with no serious adverse events in abdominal surgery. In conclusion we had better not make any traditional difference in intraoperative fluid management between abdominal and thoracic surgery even if their stress response differs in magnitude.

[Efficacy and safety of remifentanil-based regimen for postoperative pain management in abdominal surgery patients: a double-blind study with low-dose remifentanil infusion of 0.02 microg x kg(-1) x min(-1)].

BACKGROUND: Remifentanil is a powerful analgesic with fast onset and ultra-short duration of action. Its context-sensitive half-time is consistently short even after a prolonged infusion. Remifentanil is effective for providing better postoperative analgesia, but this method is not generally accepted in Japan. The present study was conducted to document efficacy and safety of low-dose remifentanil infusion in postoperative patients. METHODS: Forty patients undergoing abdominal surgery were studied prospectively. They were randomly assigned to either remifentanil (0.02 microg x kg(-1) x min(-1)) or placebo group. Postoperatively all patients received continuous epidural anesthesia with lidocaine and IV patient-controlled analgesia with fentanyl. Flurbiprofen was administered only when no pain relief was achieved. Visual analogue scale (VAS), requirement of fentanyl and flurbiprofen, and the incidence of remifentanil-related adverse effects (respiratory depression, nausea, vomiting, pruritus) were examined at 3 hourly intervals for 12 hours. RESULTS: There are no statistical differences between two groups in pain scores. No adverse events including respiratory depression occurred throughout the study in both groups. CONCLUSIONS: Remifentanil infusion at 0.02 microg x kg(-1) x min(-1) can safely be used without any serious adverse events, while it may not be enough for postoperative analgesia. The best dosage of this drug for postoperative analgesia remains to be elucidated.

[Postoperative pain management in video-assisted thoracic surgery using a continuous unilateral intercostal analgesia].

BACKGROUND: Optimal pain management after video-assisted thoracic surgery (VATS) remains an open issue. We prospectively studied the analgesic effect of intercostal analgegia (ICA) by comparison with epidural analgesia. METHODS: Twenty-two patients undergoing VATS procedures were randomly divided into ICA (n = 8) or epidural (n = 14) group. Postoperatively 2 ml x hr(-1) of 0.2% ropivacaine was delivered continuously through intercostal or epidural catheter. Moreover, each group received the equal dose of fentanyl (0.25 x µg(-1). kg(-1) x hr(-1)) intravenously or epidurally. When no pain relief was achieved, iv fentanyl was given as a rescue. Requirement of additional fentanyl and pain score using a visual analogue scale (VAS) were documented for 19 hours. RESULTS: The mean pain scores at rest, mobilization and with coghing were slightly higher in the ICA group. Total additional dose of iv fentanyl was significantly different between the groups (ICA 147 ± 41 vs Epidural 39 ± 15 µg; P = 0.015). Pain scores and fentanyl requirements spread over the lower range. The mean of VAS in ICA group was less than 5 even at coughing, suggesting clinically irrelevant. CONCLUSIONS: In patients with coagulopathy, multimodal approach using intercostal analgesia supplemented by intravenous patient-controlled analgesia may be an alternative to epidural analgesia for postoperative pain management.

[Intraperitoneal irrigation for pseudomyxoma peritonei-a case of critical metabolic alkalosis precipitated by irrigation with 101 of sodium bicarbonate--].

Pseudomyxoma peritonei causes marked accumulation of jelly-like ascites in the peritoneal cavity. Removal of much mucinous ascites by irrigating the cavity appears to be an effective treatment. We describe a patient who underwent the irrigation with sodium bicarbonate solution and developed critical alkalemia. A 68-year-old woman with normal renal function was operated on for recurrent pseudomyxoma peritonei. Fol- lowing the excision of primary lesion, her intraperitoneal cavity was irrigated with 10 1 of 7% sodium bicarbonate in about 45 minutes. Thirty minutes after irrigation, blood gas analysis revealed severe metabolic alkalosis (pH 7.714, BE 25.6 mmol x l-1 ) with electrolyte disorder (Na 157.8 mmol x l-1 K 2.31mmol x l-1, Ca 0.73 mmol x l-1). Hypotension (<60 mmHg) and sinus tachycardia (>130 beats x min -1) supervened 75 minutes later. Transferring to the ICU, she was given KC1 solution intravenously based on serial blood analysis while on mechanical ventilation. The next day acid-base disturbance returned spontaneously to normal (pH 7.45, BE 8.0mmol x l-1), leading to endotracheal extubation. Electrolyte imbalance was gradually resolved on 2nd POD and she was discharged from the ICU. Intraperitoneal irrigation with sodium bicarbonate requires special perioperative considerations for lifethreatening alkalemia, especially in a patient with renal impairment.

[The evaluation of total body water composition during caesarean section under spinal anesthesia].

BACKGROUND: Hypotension is a common adverse effect of spinal anesthesia (SA). Preoperative fluid infusion is recommended to prevent hypotension during caesarean section. The aim of this study is to document relationship between preoperative total body water (TBW) and the amount of the vasopressors given intraoperatively and to evaluate the change of maternal body water composition (BWC). METHODS: In 57 patients scheduled for caesarean section under SA, maternal BWC was measured by bioelectrical impedance analysis method. SA was performed using 0.5% isobaric bupivacaine 2 ml and fentanyl 20 microg at L3-4 intervertebral space. After this procedure, the impedance was measured again. We investigated the correlation between TBW and the amount of the vasopressor and the change of maternal BWC before and after SA. RESULTS: No positive linear correlation existed between the preoperative TBW and the amount of vasopressors given intraoperatively. SA produced a 3.8% increase in TBW and a 4.7% increase in extracellular water (ECW, P < 0.01). CONCLUSIONS: Preoperative TBW does not affect the amount of vasopressors given during caesarean section, which suggested massive fluid infusion can not always prevent hypotension after SA. A slight changes in TBW and ECW may be induced by SA, while the exact physiological and clinical significance of these observation remains to be elucidated.

[A case-control study of airway management for 68 patients with cervical spine injury: comparison of the direct laryngoscope with a Macintosh blade and the fiberoptic bronchoscope].

BACKGROUND: As for cervical spine injury, special consideration for airway management is required but the optimal strategy remains controversial. Direct laryngoscopy (DL) creates some degree of cervical extension leading to secondary neurologic deterioration. Fiberoptic bronchoscopy (FOB) may facilitate tracheal intubation with little cervical motion, but has several inherent limitations. A few objective data prompted us to compare the neurologic outcome relating to the orotracheal intubation using the different types of technique, the DL with a Macintosh blade or the FOB. METHODS: To identify the effect of different methods on the intubation time, neurologic disability, and adverse effects, 68 cervical spine-injured patients with the use of DL (group L; 36 patients) or FOB (group F; 32 patients) were retrospectively reviewed using hospital records. Following the induction of general anesthesia, the trachea was intubated with no immobilizing forces in group L, while awake intubation was accomplished in group F after judicious application of local anesthesia to the larynx and trachea. RESULTS: No significant differences were observed between the groups in age, BMI, intubation time, postoperative neurologic outcome or incidence of aspiration pneumonia. Moreover, no neurologic deterioration was shown after DL and orotracheal intubation. CONCLUSIONS: We found no evidence to support the routine practice of the bronchoscope-assisted awake intubation in patients with cervical spine injury. The clinical value of this technique in offering some neurologic advantage remains limited.

[Anesthetic considerations for spine surgery in 71 patients with infectious spondylitis: effects of different pathogen either pyogenic or tuberculous on intraoperative blood loss].

BACKGROUND: Little information is available about anesthetic management in spine surgery for infectious spondylitis, in which major bleeding can be expected. The amount of blood loss may vary somewhat with pyogenic or tuberculous spondylitis. Limited data prompted us to get a clue to determine how best to care for these patients. METHODS: To examine the amount of intraoperative bleeding, 71 patients with either pyogenic (group A; 44 patients) or tuberculous spondylitis (group B; 27 patients) were retrospectively reviewed using hospital records. They underwent posterior fusion with instrumentation and anterior radical resection of the lesion. RESULTS: No significant differences were observed between the groups in age, gender, comorbidity or length of hospital stay. Operative time was longer in patients with group B (A: 126 +/- 41 vs B: 197 +/- 76 min, P<0.01). There was a trend toward greater blood loss in group B, especially massive bleeding (>1.5 l) occurred at a higher rate (13.6 vs 33.3%, P=0.05). The number of involved vertebrae was more in group B (1.8 +/- 0.9 vs 2.9 +/- 1.3, P<0.01). Both operative time and blood loss volume showed a good correlation with the number of vertebrae infected, suggesting that extensive eradication over several spinal segments may be indicated for tuberculous spondylitis. CONCLUSIONS: Spine surgery for tuberculous spondylitis is more likely to carry risks of longer operative time and higher rate of blood loss.

[Case of obstructive sleep apnea possibly having led to postoperative appearance of generalized convulsion].

Adverse surgical outcomes appear to be more frequent in patients with known obstructive sleep apnea (OSA). However, OSA patients may present for surgery without a prior diagnosis. A 37-year-old man underwent craniotomy for surgical direct neck clipping of the right ruptured internal carotid aneurysm. His intraoperative and early postoperative courses were uneventful. At night, about 48 hr after surgery, he developed sudden generalized tonic-clonic convulsion and temporary depressed consciousness resulting in marked hypercapnea (Pa(CO2)>100 mmHg). His respiration was transiently supported by PSV mode via LMA. He soon got well without neurologic deficits. At night, about 74 hr postoperatively, a generalized convulsion was again observed with hypercapnea. Aside from the respiratory support, percutaneous cricothyroidotomy was performed using Minitrach II system for his airway control, leading to no further recurrence of seizure. He was suspected to have unrecognized OSA due to such characteristic findings of sleep apnea as obesity (BMI>30) and witnessed apneas by his family. Postoperative rapid eye movement (REM) sleep rebound has been suggested to contribute to two consecutive night appearance of seizure. Clinical suspicion for OSA should be required preoperatively and perioperative heightened awareness is recommended.

[Cardiac arrest induced by dexmedetomidine].

A 64-year-old woman with hypertension, diabetes mellitus and asymptomatic first degree AV block underwent low anterior resection of the rectum. Anesthesia was induced with propofol, vecuronium bromide and remifentanil and maintained with nitrous oxide in oxygen, propofol and remifentanil. We did not use epidural anesthesia. After the operation, the patient was admitted to the intensive care unit under general anesthesia with propofol and remifentanil. In addition, dexmedetomidine was given without loading dose. The EKG changed from first degree AV block to second degree AV block followed by complete AV block and finally cardiac arrest. As soon as we performed heart massage, sinus rhythm appeared. We should be careful in giving dexmedetomidine to a patient with AV block.

Selective loss of nigral dopamine neurons induced by overexpression of truncated human alpha-synuclein in mice.

Parkinson's disease is characterized by loss of nigral dopaminergic neurons and presence of Lewy bodies, whose major component is alpha-synuclein. In the present study, we generated transgenic mice termed Syn130m that express truncated human alpha-synuclein (amino acid residue number: 1-130) in dopaminergic neurons. Notably, dopaminergic neurons were selectively diminished in the substantia nigra pars compacta of Syn130m, while transgenic mice that expressed comparable amount of full-length human alpha-synuclein did not develop such pathology. Therefore, the truncation of human alpha-synuclein seems to be primarily responsible for the loss of nigral dopaminergic neurons. The nigral pathology resulted in impairment of axon terminals in the striatum and concomitant decrease in striatal dopamine content. Behaviorally, spontaneous locomotor activities of Syn130m were reduced, but the abnormality was ameliorated by treatment with L-DOPA. The loss of nigral dopaminergic neurons was not progressive and seemed to occur during embryogenesis along with the onset of expression of the transgene. Our results indicate that truncated human alpha-synuclein is deleterious to the development and/or survival of nigral dopaminergic neurons.

Dopamine receptor agonists reverse behavioral abnormalities of alpha-synuclein transgenic mouse, a new model of Parkinson's disease.

Parkinson's disease (PD) is characterized by loss of nigral dopaminergic (DAergic) neurons and presence of Lewy bodies, whose major component is alpha-synuclein. We had previously generated transgenic mice termed Syn130m that express truncated human alpha-synuclein (amino acid residues 1-130) in DAergic neurons. Syn130m mice showed significant loss of DAergic neurons in the substantia nigra pars compacta. Subsequently, the striatal DA level and spontaneous locomotor activity of the mice were decreased significantly. In the present study, we investigated behavioral responses of Syn130m mice to L-DOPA and DA receptor agonists. Administration of L-DOPA dose dependently ameliorated the reduction of spontaneous locomotor activity of Syn130m mice. Similarly, D(2) agonists, quinpirole and talipexole, and a D1/D2 agonist, pergolide, were effective against the reduction. Syn130m mice also showed significant reduction in exploratory behavior compared with non-Tg littermates when they were placed in a novel environment, but this abnormality was ameliorated by treatment with pergolide. These results strongly suggest that the behavioral abnormalities of Syn130m mice were caused by low striatal DA content. On the other hand, the expression of postsynaptic D(2)-like receptors (DRD2) in the striatum was not increased in Syn130m mice, although the low striatal DA level is known to induce compensatory expression of DRD2. Because the abnormalities could be rectified by treatment with DA receptor agonists, it is likely that Syn130m mice provide a useful tool to explore therapeutic possibilities for PD as a new animal model of the disease.

Accumulation of phosphorylated alpha-synuclein in dopaminergic neurons of transgenic mice that express human alpha-synuclein.

Parkinson's disease is neuropathologically characterized by the presence of Lewy bodies, whose major component is alpha-synuclein. We had previously generated transgenic mice that expressed human alpha-synuclein carrying an Ala53Thr point mutation (halpha-syn140m) under the control of the rat tyrosine hydroxylase promoter and found that halpha-syn140m was localized not only in the cytoplasm but also in the nuclei of mesencephalic dopaminergic neurons. In the present study, we carried out immunohistochemical analysis of the brain of Tg mice using anti-PSer129, an antibody that specifically recognizes alpha-synuclein phosphorylated at Ser129. The antibody detected only phosphorylated halpha-syn140m, whereas phosphorylation of endogenous alpha-synuclein, if any, was below the detection limit of the method employed. The analysis showed that approximately one-third of the halpha-syn140m-positive neurons in the midbrain of heterozygous Tg mice were concomitantly reactive to anti-PSer129. The ratio almost doubled in homozygotes, indicating that the phosphorylation level depends directly on the amount of substrate. In addition, the ratio did not change at least up to 48 weeks of age. These data strongly suggest that halpha-syn140m underwent constitutive phosphorylation and that the phosphorylation level was maintained to a certain level until the aged stages. Remarkably, halpha-syn140m localized in the nuclei seemed to be preferentially phosphorylated compared with that in the cytoplasm. Among kinases that have been reported to be involved in the phosphorylation of alpha-synuclein, the beta subunit of casein kinase-2 was detected in the nuclei by immunohistochemistry. These data imply that at least casein kinase-2 is involved in the phosphorylation of halpha-syn140m in the Tg mice.