Tomoya Okada

Ketamine inhibits monoamine transporters expressed in human embryonic kidney 293 cells.

Background Ketamine has been characterized as having psychotomimetic and sympathomimetic effects. These symptoms have raised the possibility that ketamine affects monoaminergic neurotransmission. To elucidate the relation between ketamine and monoamine transporters, the authors constructed three cell lines that stably express the norepinephrine, dopamine, and serotonin transporters and investigated the effects of ketamine on these transporters. Methods Human embryonic kidney cells were transfected using the Chen‐Okayama method with the human norepinephrine rat dopamine, and rat serotonin transporter cDNA subcloned into the eukaryotic expression vector. Using cells stably expressing these transporters, the authors investigated the effects of ketamine on the uptake of these compounds and compared them with those of pentobarbital. Results Inhibition analysis showed that ketamine significantly inhibited the uptake of all three monoamine transporters in a dose‐dependent manner. The Ki (inhibition constant) values of ketamine on the norepinephrine, dopamine, and serotonin transporters were 66.8 micro Meter, 62.9 micro Meter, and 162 micro Meter, respectively. Pentobarbital, a typical general anesthetic agent with no psychotic symptoms, did not affect the uptake of monoamines, however. Further, neither the glycine transporter 1 nor the glutamate/aspartate transporter was affected by ketamine, indicating that ketamine preferentially inhibits monoamine transporters. Conclusions Ketamine inhibited monoamine transporters expressed in human embryonic kidney cells in a dose‐dependent manner. This result suggests that the ketamine‐induced inhibition of monoamine transporters might contribute to its psychotomimetic and sympathomimetic effects through potentiating monoaminergic neurotransmission.

Comparison between the decrease of dopamine transporter and that of L-DOPA uptake for detection of early to advanced stage of Parkinson's disease in animal models.

Early diagnosis of Parkinsons disease (PD) is important for the potential application of neuroprotective therapies. The purpose of this study was to assess the detection of the early changes of PD by either imaging the dopamine transporter (DAT) or uptake of l‐3,4‐dihydroxyphenylalanine (l‐DOPA). An early to advanced stage model of PD was induced in rats by stereotaxic injection of 1–10 μg 6‐hydroxydopamine (6‐OHDA) into the substantia nigra pars compacta. Using adjacent sections of the same animals, the binding of [I‐125]beta‐CIT, which labels DAT and the uptake of [C‐14]l‐DOPA, were evaluated 4 weeks after induction of the lesion. Any decrease in dopaminergic neurons was evaluated by in situ hybridization histochemistry (ISH) by detection of DAT mRNA‐positive neurons. In addition, the expression levels of DAT, dopa decarboxylase (DDC), and vesicular monoamine transporter (VMAT2) in each neuron were studied with ISH. Our results show a decrease in both [I‐125]beta‐CIT binding and [C‐14]l‐DOPA uptake in parallel with a decrease in DA neurons from early to advanced stage models of PD. The decrease in [C‐14]l‐DOPA uptake was smaller than that in [I‐125]beta‐CIT binding in the same animal (P < 0.0001). Expression levels of DAT, DDC, and VMAT2 mRNAs were also decreased with the progression of the disease. Although ISH failed to detect the origin of the discrepancy between [I‐125]beta‐CIT and [C‐14]l‐DOPA levels, it was concluded that [C‐14]l‐DOPA levels underestimated the decrease of dopaminergic neurons and that [I‐125]beta‐CIT levels more precisely reflected the decrease. Synapse 31:178–185, 1999.

Assessment of affinities of beta-CIT, beta-CIT-FE, and beta-CIT-FP for monoamine transporters permanently expressed in cell lines

We investigated the effects of three cocaine analogs, beta-CIT (2-beta-carbomethoxy-3-beta-(4-iodophenyl)-tropane), beta-CIT-FE (2-beta-carbomethoxy-3-beta-(4-iodophenyl)-N-(2.fluoroethyl)-nortropa ne), and beta-CIT-FP (2-beta-carbomethoxy-3-beta-(4-iodophenyl)-N-(3-fluoropropyl)-n ortropane), on the uptake of [3H]dopamine(DA), serotonin(5-HT), and 1-norepinephrine (NE) using cell lines permanently expressing DA, 5-HT, and NE transporters, respectively, to determine their affinities for these three transporters. We generated cell lines stably expressing DA, 5-HT, and NE transporters, respectively, by the Chen-Okayama method, and then tested the abilities of (-)cocaine, beta-CIT, beta-CIT-FE, beta-CIT-FP, and clomipramine to inhibit the uptake of [3H]DA, 5-HT, and 2-NE. Ki values of beta-CIT, beta-CIT-FE, and beta-CIT-FP for [3H]DA, 5-HT, 1-NE uptake were 6, 29, and 33 nM, 91, 133, and 130 nM, and 28, 113 and 70 nM, respectively, whereas those of cocaine and clomipramine were 316, 581, and 176 nM and > 10,000, 437, and 851 nM, respectively. Beta-CIT, beta-CIT-FE, and beta-CIT-FP were shown to be potent DA, 5-HT, and NE uptake inhibitors. Beta-CIT and beta-CIT-FP were highly potent and selective dopamine uptake inhibitors, and therefore might be useful for imaging of DA transporter with single photon emission computed tomography (SPECT) or positron emission tomography (PET).

Detection of impaired fatty acid metabolism in right ventricular hypertrophy: Assessment by I-123 β-methyl iodophenyl pentadecanoic acid (BMIPP) myocardial single-photon emission computed tomography

Fatty acid metabolism has been reported to be impaired earlier than myocardial blood flow in left ventricular hypertrophic myocardium, e.g., in hypertrophic cardiomyopathy or hypertensive heart disease. The purpose of this study was to determine whether impaired fatty acid metabolism also occurs in right ventricular (RV) hypertrophy. The subjects consisted of 6 patients with chronic obstructive pulmonary disease, 4 with primary pulmonary hypertension, 2 each with refractory pulmonary tuberculosis, tricuspid insufficiency, pulmonary embolism, 1 each with atrial septal defect, ventricular septal defect (Eisenmenger complex), Ebstein anomaly, and endocardial defect, and 7 healthy controls. SPECT imaging with Tl-201 (Tl) and I-123 β-methyliodophenyl pentadecanoic acid (BMIPP), and Tc-99m RBC first pass and gated blood pool scintigraphy were performed. Based on Tl planar images, the subjects were classified into 3 groups: 7 patients with no RV visualization (Group A), 11 with moderate RV visualization (Group B) and 9 with marked RV visualization (Group C). As a semi-quantitative evaluation by Tl myocardial SPECT, 3 regions in 3 representative short axial images were divided into 9 segments, each of which was graded from 0 to +3, and their sum was calculated as the RV score. The right ventricular ejection fraction (RVEF) and the left ventricular ejection fraction were obtained by Tc-99m RBC cardiac scintigraphy. The groups with marked visualization of the right ventricle had lower RVEF (p < 0.01), and there was a good correlation between the RVEF and the RV score with both Tl and BMIPP (Tl: r = −0.79, BMIPP: r = −0.70). Although a good correlation was demonstrated between the RV score with Tl and BMIPP in Groups A and B (r = 0.86, p < 0.001), in Group C, in which there was marked RV Tl visualization, the RV score with BMIPP was significantly smaller than with Tl (BMIPP vs. Tl: 11.5 ± 3.7 vs. 16.4 ± 3.8, p < 0.01). These findings suggest that impaired fatty acid metabolism may exist in severely hypertrophic right ventricle due to RV overload.

MK-801 blocks monoamine transporters expressed in HEK cells

(+)‐MK‐801 is known to be a specific non‐competitive antagonist of N‐methyl‐d‐aspartate (NMDA) receptors. However, besides having an anticonvulsant effect, this compound possesses a central sympathomimetic effect and an anxiolytic‐like action, raising the possibility that (+)‐MK‐801 might affect monoamine uptake systems. To elucidate this possibility, we investigated the effects of (+)‐MK‐801 on monoamine transporters expressed in HEK cells. (+)‐MK‐801 significantly inhibited the uptake of all three monoamine transporters in a dose‐dependent manner and the inhibitions were competitive with respect to monoamines. The K i values of (+)‐MK‐801 on the norepinephrine, dopamine and serotonin transporters were 3.2 μM, 40 μM and 43 μM, respectively. In addition, (−)‐MK‐801, a less potent antagonist of NMDA receptors, also inhibited monoamine transporters with a similar potency as that of (+)‐MK‐801. These results clearly indicate that MK‐801, a non‐competitive antagonist of NMDA receptors, competitively inhibits monoamine transporters without stereoselectivity.

A new method to evaluate ischemic heart disease: combined use of rest thallium-201 myocardial SPECT and Tc-99m exercise tetrofosmin first pass and myocardial SPECT.

We developed a new diagnostic method for simultaneously evaluating myocardial ischemia, myocardial viability and ventricular function in less than 90 minutes by combined use of rest thallium-201 (Tl) SPECT and exercise Tc-99m tetrofosmin (TF) first pass and SPECT. The subjects were 9 healthy controls, 19 angina pectoris patients, and 19 old myocardial infarction patients, in all of whom coronary angiography had been performed. Rest Tl myocardial SPECT was performed first, and was followed by exercise TF myocardial SPECT. We also performed first pass radionuclide angiography by TF during maximum exercise on a bicycle ergometer to assess the left ventricular ejection fraction (LVEF). The total examination time was less than 90 minutes. SPECT diagnosis was performed by semi-quantitative analysis. LVEF below 55% was regarded as abnormal. In the patients with angina pectoris, analysis according to the coronary artery showed that the diagnostic accuracy of SPECT was 85.0% for ischemia in the region of the left anterior descending branch (LAD), 87.5% for the left circumflex branch (LCX) and 77.8% for the right coronary artery (RCA). The accuracy of diagnosis for angina pectoris was 82.1 %, as determined by SPECT alone, and rose to 89.3% when the LVEF levels were also taken into consideration. In the patients with old myocardial infarction, the diagnostic accuracy of SPECT was 84.2% for the LAD, 92.3% for the LCX and 85.0% for the RCA. Analysis by patients showed that the accuracy of diagnosis for myocardial infarction was 85.7%, as determined by SPECT alone. The diagnostic accuracy, however, rose to 89.3% when the LVEF levels also were taken into consideration. In conclusion, it was demonstrated that this combined diagnostic method was highly reliable for evaluating ischemic heart disease within a short time.

259 Ketamine inhibits monoamine transporters

Migita, keisuke ‘, Hori, Nobuaki2, Saito, Rye’ , Kenji, Yamamoto2, Talcano, Yukio’ , Kamiya, Hiro-o’ The area postrema (AP) and the nucleus tractus solitarius (NTS) play an important role for cardiovascular regulation. Our previous studies have shown that the injection of arginine vasopressin (AVP) into the AP modifies arterial baroreceptor reflex. However, it is not clear about the functional connection between the AP and the NTS. In this studies, we electrophysiologically examined the effects of AVP on the single neuron in the AP and discussed the functional relationship between the AP and NTS. Colonal and semi-horizontal slices containing the area postrema (AP) were preparad from male Wistar rats (about 250g). Stimulating electrodes were set on the AP and solitary tract (ST), and spikes were recorded from the NTS neuron. Most of neurons in the AP and NTS showed spontaneous firings, and some of neurons in the NTS received the inputs from both the AP and ST. AVP (10-s M-10m6 M) caused inhibitory responses (24.3%) and excitatory response (24.7%) in the AP neurons, respective