Time-synchronized 2-deoxy-2-[18F]fluoro-D-glucose PET/MRI with MR-active trigger and Bayesian penalized likelihood reconstruction: Diagnostic utility for locoregional extension of endometrial cancer.

PURPOSE: Minimal misregistration of fused PET and MRI images can be achieved with simultaneous positron emission tomography/magnetic resonance imaging (PET/MRI). However, the acquisition of multiple MRI sequences during a single PET emission scan may impair fusion precision of each sequence. This study evaluated the diagnostic utility of time-synchronized PET/MRI using an MR active trigger and a Bayesian penalized likelihood reconstruction algorithm (BPL) to assess the locoregional extension of endometrial cancer. METHODS: Fifty-five patients with endometrial cancer who underwent pelvic 2-deoxy-2-[18F]fluoro-D-glucose PET/MRI were retrospectively evaluated. The PET emission time for the BPL reconstruction was determined by the MR active trigger of each MR sequence. The concordance rates of image interpretation with pathological T-staging, diagnostic performance for deep myometrial invasion (MI), and diagnostic confidence levels were evaluated by two readers and compared between time-synchronized, overlapping (conventional and simultaneous, but not time-synchronized), and sequential (not simultaneous) PET/MRI and MRI with diffusion-weighted imaging. Misregistration of the PET/MRI-fused images was determined by evaluating the differences in bladder dimensions. RESULTS: The T classification by time-synchronized PET/MRI was the most concordant with the pathological T classification for the two readers. Time-synchronized PET/MRI had a significantly higher diagnostic performance for deep MI and higher confidence level scores than overlapping PET/MRI for the novice reader (p = 0.033 and p = 0.038, respectively). The differences in bladder dimension on sequential PET/MRI were significantly larger than those on overlapping and time-synchronized PET/MRI (p <0.001). CONCLUSION: Time-synchronized PET/MRI is superior to conventional PET/MRI for assessing the locoregional extension of endometrial cancer.

Nano-Tactile Scanner with Dust-Proof and Drip-Proof Structure for High-Resolution Measurement of Skin Surface Textures.

In this study, we have successfully developed a scanner-type measurement device with a built-in nano-tactile sensor for measuring living organisms and skin surface. A complete sensor protection structure for sensitive tactile sensor device has been developed to achieve a sensing system that is capable of measuring texture changes on the skin surface. By using a highly flexible organic ultra-thin film (film adhesive bandage) as the protective structure, it is possible to prevent deterioration of sensor performance and the intrusion of droplets and dust present on the surface to be measured. This system was used in a clinical experiment at the university's medical faculty, and succeeded in accurately extracting changes in the properties of the patient's skin caused by different wiping methods. As a result, the measurement of tactile texture of the skin surface in various states between dry and wet could be accurately done. Finally, in addition to the ability to measure tactile characteristics of the skin surface, a new function has been realized to measure skin hardness distribution caused by skin changes due to blisters, moles, etc.Clinical Relevance- The developed micron scale was the first to explain the changes in the skin surface structure according to the type of skin surface stimulus and the time of its appearance. It is clinically useful to interpret the protective function of the skin and the function of sensory reception.

Rapid Whole-Body FDG PET/MRI in Oncology Patients: Utility of Combining Bayesian Penalised Likelihood PET Reconstruction and Abbreviated MRI.

This study evaluated the diagnostic value of a rapid whole-body fluorodeoxyglucose (FDG) positron emission tomography (PET)/magnetic resonance imaging (MRI) approach, combining Bayesian penalised likelihood (BPL) PET with an optimised ß value and abbreviated MRI (abb-MRI). The study compares the diagnostic performance of this approach with the standard PET/MRI that utilises ordered subsets expectation maximisation (OSEM) PET and standard MRI (std-MRI). The optimal ß value was determined by evaluating the noise-equivalent count (NEC) phantom, background variability, contrast recovery, recovery coefficient, and visual scores (VS) for OSEM and BPL with ß100-1000 at 2.5-, 1.5-, and 1.0-min scans, respectively. Clinical evaluations were conducted for NECpatient, NECdensity, liver signal-to-noise ratio (SNR), lesion maximum standardised uptake value, lesion signal-to-background ratio, lesion SNR, and VS in 49 patients. The diagnostic performance of BPL/abb-MRI was retrospectively assessed for lesion detection and differentiation in 156 patients using VS. The optimal ß values were ß600 for a 1.5-min scan and ß700 for a 1.0-min scan. BPL/abb-MRI at these ß values was equivalent to OSEM/std-MRI for a 2.5-min scan. By combining BPL with optimal ß and abb-MRI, rapid whole-body PET/MRI could be achieved in ≤1.5 min per bed position, while maintaining comparable diagnostic performance to standard PET/MRI.

PMEL p.Leu18del dilutes coat color of Kumamoto sub-breed of Japanese Brown cattle.

BACKGROUND: Coat color is important for registration and maintenance of livestock. Standard coat color of Kumamoto sub-breed of Japanese Brown cattle is solid brown, but individuals with diluted coat color have been observed recently. In this study, we attempted to identify polymorphism(s) responsible for coat color dilution by whole genome analysis. RESULTS: One of the diluted cattle possessed 7302 exonic polymorphisms which could affect genes' function. Among them, 14 polymorphisms in 10 coat color-related genes were assumed to be specific for the diluted cattle. Subsequent genotyping with three diluted cattle and 74 standard cattle elucidated that PMEL p.Leu18del was the causative polymorphism for coat color dilution in this sub-breed. Individuals with del/del type of this polymorphism showed diluted coat color, but coat color of heterozygotes were intermediate with various dilution rates. CONCLUSIONS: Coat color dilution of Kumamoto sub-breed was caused by PMEL p.Leu18del. The causative del allele has been detected in several genetically distant cattle breeds, suggesting that PMEL p.Leu18del can be used as a DNA marker to control cattle coat color.

Zeolite Improves High-Fat Diet-Induced Hyperglycemia, Hyperlipidemia and Obesity in Mice.

Zeolite, an abundant mineral in the Earth's crust, is utilized in a wide range of fields because of its well-known adsorption properties. Its application as a functional food ingredient resembling dietary fiber is expected, but it has not yet been investigated in the context of prevention of lifestyle-related diseases. The present study was designed to evaluate the availability and safety of a natural zeolite preparation for this purpose. Acute oral toxicity testing showed that the lowest lethal dose of zeolite was more than 2,000 mg/kg body weight for both male and female mice. In a prolonged feeding test for 18 wk using model mice with high-fat-induced obesity and type 2 diabetes mellitus, intake of a 10% zeolite-containing diet suppressed body weight gain, as well as liver and visceral fat weights, without any changes in food and energy intake. Moreover, plasma lipid (triacylglycerol, total cholesterol and high-density-lipoprotein cholesterol) levels and fasting blood glucose levels decreased in parallel with zeolite intake. No changes in the glycated hemoglobin level were found. However, in an oral glucose tolerance test at week 12, increased postprandial blood glucose levels were suppressed in accordance with zeolite intake, and then insulin secretion was also decreased. On the other hand, a decrease of plasma amylase activity and increases in total bilirubin and urea nitrogen suggested the need for further investigation of safety.

Physiological skin FDG uptake: A quantitative and regional distribution assessment using PET/MRI.

PURPOSE: To retrospectively assess the repeatability of physiological F-18 labeled fluorodeoxyglucose (FDG) uptake in the skin on positron emission tomography/magnetic resonance imaging (PET/MRI) and explore its regional distribution and relationship with sex and age. METHODS: Out of 562 examinations with normal FDG distribution on whole-body PET/MRI, 74 repeated examinations were evaluated to assess the repeatability and regional distribution of physiological skin uptake. Furthermore, 224 examinations were evaluated to compare differences in the uptake due to sex and age. Skin segmentation on PET was performed as body-surface contouring on an MR-based attenuation correction map using an off-line reconstruction software. Bland-Altman plots were created for the repeatability assessment. Kruskal-Wallis test was performed to compare the maximum standardized uptake value (SUVmax) with regional distribution, age, and sex. RESULTS: The limits of agreement for the difference in SUVmean and SUVmax of the skin were less than 30%. The highest SUVmax was observed in the face (3.09±1.04), followed by the scalp (2.07±0.53). The SUVmax in the face of boys aged 0-9 years and 10-20 years (1.33±0.64 and 2.05±1.00, respectively) and girls aged 0-9 years (0.98±0.38) was significantly lower than that of men aged ≥20 years and girls aged ≥10 years (p<0.001). In women, the SUVmax of the face (2.31±0.71) of ≥70-year-olds was significantly lower than that of 30-39-year-olds (3.83±0.82) (p<0.05). CONCLUSION: PET/MRI enabled the quantitative analysis of skin FDG uptake with repeatability. The degree of physiological FDG uptake in the skin was the highest in the face and varied between sexes. Although attention to differences in body habitus between age groups is needed, skin FDG uptake also depended on age.

Diagnostic performance of zero-TE lung MR imaging in FDG PET/MRI for pulmonary malignancies.

OBJECTIVES: This study aimed to evaluate the diagnostic performance of the lung zero-echo time (ZTE) sequence in FDG PET/MRI for detection and differentiation of lung lesions in oncologic patients in comparison with conventional two-point Dixon-based MR imaging. METHODS: In this single-institution retrospective study approved by the institutional review board, 209 patients with malignancies (97 men and 112 women; age range, 17-89 years; mean age, 66.5 ± 12.9 years) underwent 18F-FDG PET/MRI between August 2017 and August 2018, with diagnostic Dixon and ZTE under respiratory gating acquired simultaneously with PET. Image analysis was performed for PET/Dixon and PET/ZTE fused images by two readers to assess the detectability and differentiation of lung lesions. The reference standard was pathological findings and/or the data from a chest CT. The detection and differentiation abilities were evaluated for all lesions and subgroups divided by lesion size and maximum standardized uptake value (SUVmax). RESULTS: Based on the reference standard, 227 lung lesions were identified in 113 patients. The detectability of PET/ZTE was significantly better than that of PET/Dixon for overall lesions, lesions with a SUVmax less than 3.0 and lesions smaller than 4 mm (p < 0.01). The diagnostic performance of PET/ZTE was significantly better than that of PET/Dixon for overall lesions and lesions smaller than 4 mm (p < 0.01). CONCLUSIONS: ZTE can improve diagnostic performance in the detection and differentiation of both FDG-avid and non-FDG-avid lung lesions smaller than 4 mm in size, yielding a promising tool to enhance the utility of FDG PET/MRI in oncology patients with lung lesions. KEY POINTS: • The detection rate of PET/ZTE for lesions with a SUVmax of less than 1.0 was significantly better than that of PET/Dixon. • The performance for differentiation of PET/ZTE for lesions that were even smaller than 4 mm in size were significantly better than that of PET/Dixon. • Inter-rater agreement of PET/ZTE for the differentiation of lesions less than 4 mm in size was substantial and better than that of PET/Dixon.

Kicked Hall systems: Quantum-dynamical and spectral manifestations of generic superweak chaos.

Classical "kicked Hall systems" (KHSs), i.e., periodically kicked charges in the presence of uniform magnetic and electric fields that are perpendicular to each other and to the kicking direction, have been introduced and studied recently. It was shown that KHSs exhibit, under generic conditions, the phenomenon of "superweak chaos" (SWC), i.e., for small kick strength κ a KHS behaves as if this strength were effectively κ^{2} rather than κ. Here we investigate quantum-dynamical and spectral manifestations of this generic SWC. We first derive general expressions for quantum effective Hamiltonians for the KHSs. We then show that the phenomenon of quantum antiresonance (QAR), i.e., "frozen" quantum dynamics with flat quasienergy (QE) bands, takes place for integer values of a scaled Planck constant ℏ_{s} and under the same generic conditions for SWC. This appears to be the most generic occurrence of QAR in quantum systems. The vicinity of QAR is shown to correspond semiclassically to SWC. A global spectral manifestation of SWC is the fact that a scaled QE spectrum as function of ℏ_{s}, at fixed small value of κ/ℏ_{s}, features an approximately "doubled" structure. In the case of standard (cosine) potentials, this structure is that of a universal (parameters-independent) double Hofstadter butterfly. Also, for standard potentials and for small ℏ_{s} (semiclassical regime), the evolution of the kinetic-energy expectation value exhibits a relatively slow quantum-diffusive behavior having universal features. These approximate spectral and quantum-dynamical universalities agree with predictions from the effective Hamiltonian.

Combined Effects of a Dietary Fiber Mixture and Wheat Albumin in a Rat Model of Type 2 Diabetes Mellitus.

As bioactive ingredients of functional foods, dietary fiber and wheat albumin (WA) are known to suppress hyperglycemia in patients with type 2 diabetes mellitus. The combined effects of these bioactive ingredients were examined using an animal model of type 2 diabetes mellitus. First, oral starch tolerance tests (OSTTs) with the simultaneous intake of a dietary fiber mixture (DF) and WA were performed as an acute study. Male Goto-Kakizaki rats received a soluble starch solution [700 mg/kg body weight (bw)] containing DF and/or WA (each 300 mg/kg bw). In these OSTTs, the combined intake of DF and WA suppressed hyperglycemia much more effectively than each separate intake. Second, in a chronic intake study, diets containing DF and/or WA were administered to male Zucker diabetic fatty rats over 84 d. The combined effects of DF and WA were not observed in glycosylated hemoglobin concentration levels or fasting blood glucose levels, but appeared as an improvement in liver lipid contents. Variations in the liver lipid contents were similarly reflected in those of the plasma lipid concentrations. In conclusion, this study found that the simultaneous intake of bioactive DF and WA improved the postprandial hyperglycemia and the chronic lipid metabolism disorders in rat models of type 2 diabetes mellitus.

Preoperative risk stratification using metabolic parameters of (18)F-FDG PET/CT in patients with endometrial cancer.

PURPOSE: To evaluate the usefulness of metabolic parameters obtained by (18)F-FDG PET/CT for preoperative stratification of high-risk and low-risk endometrial carcinomas. METHODS: Preoperative (18)F-FDG PET/CT was performed in 56 women with endometrial cancer. Maximum standardized uptake values (SUVmax), metabolic tumour volume (MTV) and total lesion glycolysis (TLG) of primary tumours were compared with clinicopathological features of surgical specimens. Diagnostic performance in terms of differentiation of low-risk disease (endometrioid histology, histological grade 1 or 2, invasion of less than half of the myometrium, and FIGO stage I) from high-risk disease was assessed. RESULTS: MTV and TLG were significantly higher in patients with higher histological grade (p = 0.0026 and p = 0.034), larger tumour size (p = 0.002 and p = 0.0017), lymphovascular space involvement (LVSI; p = 0.012 and p = 0.0051), myometrial invasion (p = 0.027 and p = 0.031), cervical stromal invasion (p = 0.023 and p = 0.014), ovarian metastasis (p = 0.00022 and p = 0.00034), lymph node metastasis (p < 0.0001 and p < 0.0001), and higher FIGO stage (p = 0.0011 and p = 0.00048). SUVmax was significantly higher in patients with larger tumour size (p = 0.0025), LVSI (p = 0.00023) and myometrial invasion (p < 0.0001). The areas under the ROC curves (AUCs) for distinguishing high-risk from low-risk carcinoma were 0.625, 0.829 and 0.797 for SUVmax, MTV and TLG, respectively. AUCs for both MTV and TLG were significantly larger than that for SUVmax (p = 0.0049 and p = 0.021). The optimal TLG cut-off value of 70.2, determined by ROC analysis, was found to have 72.0% sensitivity and 74.2% specificity for risk stratification. CONCLUSION: MTV and TLG of primary endometrial cancer show better correlations with clinicopathological features and are more useful for differentiating high-risk from low-risk carcinoma than SUVmax.

Neutralized nanoparticle composed of SS-cleavable and pH-activated lipid-like material as a long-lasting and liver-specific gene delivery system.

Charge-neutralized lipid envelope-type nanoparticles formed with SS-cleavable and pH-activated lipid-like materials (ssPalm) accumulate rapidly in the liver without forming aggregates in the blood circulation, and result in a liver-specific gene expression for a long duration (>2 weeks) with neither immunological responses nor hepatotoxicity after intraveneous administration, when it carries pDNA free from CpG-motifs.

Neutral biodegradable lipid-envelope-type nanoparticle using vitamin A-Scaffold for nuclear targeting of plasmid DNA.

Biomembranes and cytoplasm, a diffusion-limited region for nanoparticles are critical barriers to be overcome for the successful gene delivery. We herein report on a neutral, and intracellularly degradable lipid nanoparticle (LNP), containing encapsulated plasmid DNA (pDNA) that can be effectively delivered to the nucleus. A key material component in this particle is a vitamin A-scaffold SS-cleavable Proton-Activated Lipid-like Material ((SS)PalmA), which contains tertiary amine groups as proton sponge units that can respond to the acidic pH in endosomes, disulfide bonding for programmed collapse in the cytoplasm, and retinoic acid (RA) as a hydrophobic unit for assembly into LNP. LNP prepared using (SS)PalmA (LNP(PalmA)) exhibited a 15-fold higher gene expression activity compared to particles prepared with a simple acyl chain (myristoyl group)-scaffold one (LNPPalmM). Intracellular imaging studies revealed that LNP(PalmA) unexpectedly showed excessive endosome-disruptive characteristics. Furthermore, the decapsulation of pDNA slowly, but successively occurred in parallel with peri-nuclear accumulation. Nuclear targeting was blocked in the presence of native RA. Collectively, LNP(PalmA) is an intelligent particle that passes through the cytoplasm in particle form with the aid of the intrinsic nuclear transport system of RA, and thereafter releases its encapsulated pDNA for effective gene expression.

The application of pH-sensitive polymer-lipids to antigen delivery for cancer immunotherapy.

For production of pH-sensitive liposomes, we developed pH-sensitive polymer-lipids that consists of pH-sensitive fusogenic polymer moieties such as 3-methyl glutarylated poly(glycidol) and 2-carboxycyclohexane-1-carboxylated poly(glycidol), connected to a phosphatidylethanolamine head group. Incorporation of these pH-sensitive polymer-lipids into egg yolk phosphatidylcholine liposomes produced highly pH-sensitive liposomes that were stable at neutral pH but which destabilized markedly in response to very small pH change in weakly acidic pH region. These liposomes delivered their contents (pyranine) into cytosol of dendritic cell-derived DC2.4 cells. When these polymer-lipid-incorporated liposomes loaded with antigenic protein ovalbumin (OVA) were administered subcutaneously to mice, the antigen-specific cellular immunity was induced efficiently in the mice. Furthermore, immunization of mice with these OVA-loaded pH-sensitive polymer-lipid-incorporated liposomes induced strong OVA-specific immunity, which achieved complete rejection of OVA-expressing E.G7-OVA cells and marked regression of E.G7-OVA tumors.

A neutral envelope-type nanoparticle containing pH-responsive and SS-cleavable lipid-like material as a carrier for plasmid DNA.

SS-cleavable proton-activated lipid-like material (ssPalm) functions as a key element in a lipid nanoparticle in which pDNA is encapsulated. The ssPalm contains dual sensing motifs that can respond to the intracellular environment; a proton-sponge unit (tertiary amines) that functions in response to an acidic environment (endosome/lysosome), and disulfide bonding that can be cleaved in a reducing environment (cytosol).

Impaired glycinergic synaptic transmission and enhanced inflammatory pain in mice with reduced expression of vesicular GABA transporter (VGAT).

Loading of GABA and glycine into synaptic vesicles via the vesicular GABA transporter (VGAT) is an essential step in inhibitory neurotransmission. As a result of the evidence linking alterations in GABAergic and/or glycinergic neurotransmission to various pain disorders, we investigated the possible influence of down-regulation of VGAT on pain threshold and behavioral responses in mice. The phenotypes of heterozygous VGAT knockout [VGAT(+/-)] mice were compared with wild-type (WT) mice using behavioral assays. In addition, GABAergic and glycinergic miniature inhibitory postsynaptic currents (mIPSCs) were recorded in dorsal horn neurons. Western blot analysis confirmed significant reduction of VGAT protein levels in VGAT(+/-) mice. However, high-performance liquid chromatography revealed that glutamate, GABA, and glycine contents in the whole brain and spinal cord were normal in VGAT(+/-) mice. Behavioral analysis of VGAT(+/-) mice showed unchanged motor coordination, anxiety, memory performance, and anesthetic sensitivity to propofol and ketamine, although thermal nociception and inflammatory pain were enhanced. Patch-clamp recordings revealed that the frequency and amplitude of glycinergic mIPSCs in lamina II neurons were reduced in VGAT(+/-) mice. Genotype differences in glycinergic mIPSCs were more evident during sustained stimulation by solutions with high potassium levels, suggesting that the estimated size of the readily releasable pool of glycine-containing vesicles was reduced in VGAT(+/-) mice. These results provide genetic, behavioral, and electrophysiological evidence that VGAT-mediated inhibitory drive alters very specific forms of sensory processing: those related to pain processing. More close examination will be needed to verify the possibility of VGAT as a new therapeutic target for the treatment of inflammatory pain.

The influence of manipulations to alter ambient GABA concentrations on the hypnotic and immobilizing actions produced by sevoflurane, propofol, and midazolam.

Recent studies have suggested that extrasynaptic GABA(A) receptors, which contribute tonic conductance, are important targets for general anesthetics. We tested the hypothesis that manipulations designed to alter ambient GABA concentrations (tonic conductance) would affect hypnotic (as indicated by loss of righting reflex, LORR) and immobilizing (as indicated by loss of tail-pinch withdrawal reflex, LTWR) actions of sevoflurane, propofol, and midazolam. Two manipulations studied were 1) the genetic absence of glutamate decarboxylase (GAD) 65 gene (GAD65-/-), which purportedly reduced ambient GABA concentrations, and 2) the pharmacological manipulation of GABA uptake using GABA transporter inhibitor (NO-711). The influence of these manipulations on cellular and behavioral responses to the anesthetics was studied using behavioral and electrophysiological assays. HPLC revealed that GABA levels in GAD65-/- mice were reduced in the brain (76.7% of WT) and spinal cord (68.5% of WT). GAD65-/- mice showed a significant reduction in the duration of LORR and LTWR produced by propofol and midazolam, but not sevoflurane. NO-711 (3 mg/kg, ip) enhanced the duration of LORR and LTWR by propofol and midazolam, but not sevoflurane. Patch-clamp recordings revealed that sevoflurane (0.23 mM) slightly enhanced the amplitude of tonic GABA current in the frontal cortical neurons; however, these effects were not strong enough to alter discharge properties of cortical neurons. These results demonstrate that ambient GABA concentration is an important determinant of the hypnotic and immobilizing actions of propofol and midazolam in mice, whereas manipulations of ambient GABA concentrations minimally alter cellular and behavioral responses to sevoflurane.

Compensatory expression of MRP3 in the livers of MRP2-deficient EHBRs is promoted by DHA intake.

We have hypothesized a suppressive mechanism against dietary docosahexaenoic acid (22:6n-3; DHA)-induced tissue lipid peroxidation, in which the degradation products, including their conjugates, are excreted into the urine by xenobiotic or organic anion transporters. In this study, we employed parent-strain Sprague-Dawley rats (SDRs), together with their mutant strain, Eisai hyperbilirubinuria rats (EHBRs). EHBRs are deficient in multidrug resistance-associated protein (MRP) 2, and show defective urinary excretion of numerous xenobiotics and organic anions. Both strains of rats were fed a diet containing DHA at 8.4% of total energy for 31 d. In the livers of the DHA-fed rats, the level of free malondialdehyde (MDA) + 4-hydroxy-2-alkenals (HAE) fell, and conversely glutathione S-transferase (GST) activity increased in MRP2-deficient EHBRs as compared to the SDRs, suggesting that the glutathione (GSH)-conjugation reaction for the aldehydes generated on DHA intake was accelerated in the MRP2-deficient EHBRs. Since the gene expression of liver MRP3 in the MRP2-deficient EHBRs was amplified to compensate for DHA intake, it is thought that the transport of MRP3 substrates into the bloodstream, rather than MRP2-mediated excretion of its substrates into the bile, was promoted. Indeed, excretion of mercapturic acid (acetylcysteine conjugates derived metabolically from the conjugate of each aldehyde with GSH) into the urine increased significantly in MRP2-deficient EHBRs fed DHA.

Thermal hyperalgesia via supraspinal mechanisms in mice lacking glutamate decarboxylase 65.

Gamma-aminobutyric acid, which is synthesized by two isoforms of glutamate decarboxylase (GAD), inhibits the transfer of nociceptive signals from primary afferent fibers to the central nervous system. However, the roles of a 65-kDa isoform of GAD (GAD65)-mediated GABA in nociceptive processing are less clear. This study tested whether partial reductions in GABAergic inhibitory tone by GAD65 gene knockout [GAD65(-/-)] would contribute to the regulation of pain threshold in mice. Experiments were performed on male wild-type (WT) mice and GAD65(-/-) mice. Acute nociception and inflammatory pain tests were compared between WT mice and GAD65(-/-) mice. GABA(A) receptor-mediated inhibitory postsynaptic currents were also examined by use of the whole-cell patch-clamp method in somatosensory cortical neurons in brain slices. In the hot plate test, which reflects supraspinal sensory integration, a significant reduction in the latency was observed for GAD65(-/-) mice. Intraperitoneal administration of the GABA transporter 1 inhibitor, 1-[2-[[(diphenylmethylene)imino]oxy]ethyl]-1,2,5,6-tetrahydro-3-pyridinecarboxylic acid hydrochloride (C(21)H(22)N(2)O(3).HCl; NO-711), dose-dependently prolonged the latency in both genotypes, suggesting that GABA concentration contributes to acute thermal nociception. However, there was no genotype difference in responses to the tail-immersion test or the von Frey test, indicating that spinal reflex and mechanical nociception are kept intact in GAD65(-/-) mice. There was no genotype difference in responses to chemical inflammatory nociception (formalin test and carrageenan test). Although properties of the phasic component of inhibitory postsynaptic currents were similar in both genotypes, tonic inhibition was significantly reduced in GAD65(-/-) mice. These results support the hypothesis that GAD65-mediated GABA synthesis plays relatively small but significant roles in nociceptive processing via supraspinal mechanisms.

Altered responses to propofol, but not ketamine, in mice deficient in the 65-kilodalton isoform of glutamate decarboxylase.

GABA is synthesized by two isoforms of glutamate decarboxylase (GAD), GAD65, and GAD67. However, the relative contributions of GAD65-mediated GABA synthesis to the in vivo actions of anesthetics remain unknown. To address this issue, we used mice deficient in the 65-kDa isoform of GAD and tested the hypothesis that partial reduction of GABA content in GAD65-deficient mice [GAD65(-/-)] would contribute to hypnotic and immobilizing actions of the anesthetics. The open field test, loss of righting reflex (LORR), loss of tail-pinch withdrawal response (LTWR), and locomotor activity were compared between wild-type (WT) mice and GAD65(-/-) mice. Effects of general anesthetics on both phasic and tonic GABAergic currents were examined using the patch-clamp method in frontal cortex pyramidal neurons in brain slices. The duration of propofol (100 mg/kg i.p.)-induced LORR and the duration of propofol (150 mg/kg i.p.)-induced LTWR in GAD65(-/-) mice were significantly reduced compared with WT mice. In contrast, no difference was seen for ketamine. Preinjection of the GABA transporter 1 inhibitor, NO-711 (C(21)H(22)N(2)O(3).HCl) (0.75 mg/kg i.p.), reinstated diminished actions of propofol in GAD65(-/-) mice. Cortical pyramidal neurons in GAD65(-/-) mice had smaller tonic conductances, and propofol-induced enhancement of tonic inhibition was smaller than in WT mice, suggesting that genotype differences in GAD65-mediated GABAergic inhibitory tone may be, at least in part, a cellular basis underlying behavioral differences. In conclusion, GAD65(-/-) mice show a diminished response to propofol, but not ketamine, indicating that GAD65-mediated GABA synthesis plays an important role in hypnotic and immobilizing actions of propofol.