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Objective Among different PET tracers, 18F-fludeoxyglucose (FDG) and 11C-choline are known to have a high tumor uptake correlated with a high mitotic index of tumor cells. Thus, the uptake of 18F-FDG and 11C-choline may be dependent on the cell cycle. In the present study, we examined the uptake of 18F-FDG and 11C-choline in cancer cell lines by cell cycle synchronization to clarify the biological properties of cancer cells with respect to each tracer. Methods HeLa S3 Cells were synchronized by the double thymidine (TdR) block methods. 18F-FDG and 11C-choline were administered to synchronized cells, and the radioactivity per cell was measured to compare the cellular uptake of the tracers during S, G2/M, and G1 phases. Flow cytometry (FCM) was performed to measure the proportion of cells in G1, S, and G2/M phases. Furthermore, the levels of glucose transporter 1 (GLUT1) and choline transporter-like protein 1 (CTL1) in the cell were evaluated by FCM. Results The uptake of 18F-FDG was the highest in S to G2/M phases, and markedly decreased in G1 phase. The uptake of 11C-choline reached its peak in G2/M, and decreased in G1 phase. The level of GLUT1 expression was similar to that of 18F-FDG uptake during the cell cycle, and the level of CTL1 expression was similar to that of 11C-choline uptake throughout the cell cycle. Conclusions In this in vitro study, we demonstrated that 18F-FDG and 11C-choline had the highest uptake in S to G2/M phases and in G2/M phase, respectively, with a rapid decrease in G1 phase. These findings suggest that 18F-FDG and 11C-choline have a high accumulation in tumor cells with a high mitotic index. Furthermore, our study suggests that the expression of GLUT1 and CTL1 has cell cycle dependence, and the changes of 18F-FDG and 11C-choline accumulation seem to be caused by the above properties of these transporters.

Objective: Research on FDG-uptake by blood cells has revealed that FDG is incorporated by macrophages and granulocytes, as well as activated lymphocytes. These characteristics of FDG suggest the possibility of visualizing the distribution of immunocytes in target organs. The aim of this study was to investigate if mouse spleen-derived lymphocytes, activated by macrophages presenting sheep red blood cell (sRBC) antigens, could be traced by FDG.Methods: One percent of a sRBC suspension was injected into the peritoneal cavity of mice thereby creating immunity to the sRBC antigen. The splenocytes, consisting mostly of lymphocytes, were isolated, and serum containing the anti-sRBC antibody was mixed with sRBC to prepare sRBC-antibody complexes (sRBC-AbCs). Then five percent of a thioglycolate medium was injected into the peritoneal cavity of the same mice, and macrophages of ascitic cell origin were obtained. These macrophages were added to the sRBC-AbCs to induce sRBC antigen presenting macrophages. These were incubated with splenocytes obtained from sRBC immunized mouse (sRBC immunized splenocytes) or nonimmunized splenocytes to induce a T cell immune response. [3H]deoxyglucose ([3H]DG) and FDG were incorporated in splenocytes, and the quantity of their uptake was measured.Results: [3H]DG uptake by sRBC-immunized splenocytes was about eleven times as high as that of non-immunized splenocytes. In contrast, [3H]DG uptake by sRBC-immunized splenocytes, co-cultured with macrophages phagocytizing sRBC-AbCs, was about 40 times higher compared with non-immunized splenocytes. Splenocytes in non-immunized mice picked up very little [3H]DG, despite co-culture with macrophages phagocytizing sRBC-AbCs. Similar tendencies were observed with FDG.Conclusions: These results suggest that the SUV calculated in PET reflects not only the number of lymphocytes, but also the activation state of the lymphocytes themselves. In addition, the biodistribution of antigen specific lymphocytes, that have been taken up FDGin vitro and returned to the body, can be observed through PET.

Madin-Darby canine kidney (MDCK) cells were originally anchorage-dependent epithelial cells. Here, we have isolated a novel MDCK-derived cell population, termed 6 M-4, by means of culturing MDCK cells in suspension for nearly 6 months in the presence of Streptomyces griseus metalloendopeptidase (MEP). The isolated cells showed unique proliferation characteristics, which differed from parental MDCK cells. They proliferated adherently on a polystyrene matrix, but proliferated non-adherently both in the presence of MEP and on a non-adhesive matrix coated with poly 2-methacryloyloxyethyl phosphorylcholine (MPC). The 6 M-4 cells consisted of at least two cell types. One type, termed 6 M-4-TR7, would not grow in soft agar and showed a novel phenotype in that the cells were susceptible to both TNF-alpha and verotoxin 1 (VT1). In addition, the isolated adhesion-independent cells sustained epithelial traits of parental MDCK cells. We further show that these MDCK-derivative cells are suitable for influenza virus cultivation. Hemagglutination (HA) titers of influenzaviruses A and B were increased in the suspension culture of 6 M-4-TR7 cells supplemented with the MEP in comparison to adherently growing cells in the presence of trypsin.

Skinned nerve fiber was prepared by treating a giant axon of squid with saponin. Filamentous networks and membranous organelles such as vacuoles, smooth membranes and mitochondria were present after the extraction. Ruthenium red staining of the extracted tissue showed dense fibrillar networks in the cytoplasm that correspond to the "microtrabeculae" or "cytoskeleton" reported by other investigators. Our method of preparation, thus, gives further evidence that these structures are not the result of artifactual condensation or precipitation. The present results also indicated that the networks are composed of acid mucopolysaccharides and/or mucoproteins. The density of microtubules was not changed by the extraction per se, but the number of microtubules remaining after extraction decreased by alteration of environmental conditions such as low temperature (4 degrees C), or the presence of Ca++ (1 mM), Mg++ free medium or colchicine (1 mM). These conditions are known to inhibit axonal transport. The usefulness of our extracted cytoplasm as a model of microtubule-related physiological functions is discussed.

The abdominal ganglion of Aplysia includes neurons with a characteristic dopamine (DA) receptor, the activation of which induces a marked hyperpolarization with a specific increase in the permeability of the membrane to K+. The DA receptor of this type is called the "HK-type." A 2-min exposure to 1 microM serotonin (5-HT) had little effect on the resting membranes with the receptor of HK-type, but significantly depressed the responses to 10 microM DA. The depressing effect of 5-HT on this type of response was completely reversible after a 15-min washing with normal artificial Aplysia blood. Lineweaver-Burke type plotting of the DA-induced responses showed a systematic shift of the straight lines when the concentration of 5-HT was increased; the slope of the line became steeper but the intercept on the ordinate remained unchanged. The dose-inhibition curves, in which relative responses to a given [DA] were plotted against log [5-HT], showed a parallel shift toward the right when the concentration of DA increased. These findings suggest that 5-HT competes with DA for common binding sites at the DA receptor of HK-type, and that the blockade is not due to the interaction of 5-HT with K+-channels in the receptor membrane. The effect of other indole derivatives suggests that the DA receptor of HK-type includes anionic and cationic sites to which the NH2 group and 5-HO group of 5-HT could specifically bind, thus exhibiting competitive blockade.

Using the abdominal ganglion cells of Aplysia, we analyzed the effects of various enzymes and chemical modification reagents on the acetylcholine (ACh)-induced responses of the excitatory (Na+ -dependent) and inhibitory (Cl- -dependent) types. (1) Phospholipase A (2 mg/ml) caused no appreciable effects on either type of response. (2) Phospholipase C (2 mg/ml) markedly depressed both types of response. These suggested that the phosphoryl group of the phospholipid is an important site related to the binding of ACh, common to both types of ACh-receptors. (3) Carboxypeptidase A (10 mg/ml) caused no observable effects on either type of response. (4) Carboxypeptidase B (10 mg/ml) depressed the inhibitory type of response without affecting the excitatory one. (5) Pyridoxal-5'-phosphate (1 mM) also depressed the inhibitory response without affecting the excitatory one. These findings (4, 5) suggested that the Cl- -channel in the inhibitory ACh-receptor complex includes a C-terminal lysine which may play an active role in the movement of Cl- across the receptor membrane. (6) L-Leucine aminopeptidase (1 mg/ml) depressed the excitatory response without altering the inhibitory one. (7) p-Nitrothiophenol (1 mM)-also depressed the excitatory response without affecting the inhibitory one. These findings (6, 7) suggested the presence of a certain N-terminal amino acid near a glutamate or aspartate residue within a molecular moiety of Na+ -channel included in the excitatory ACh-receptor complex.

There are many cells in the abdominal ganglion which show a fast, Cl(-)-dependent hyperpolarizing response to gamma-aminobutyric acid (GABA). This response is characterized by an initial rapid increase in membrane conductance followed by an exponential decay to the original value despite a sustained application of GABA. The decay of the response was found to be largely due to the desensitization of the GABA receptor (binding site-ionophore complex) since the equilibrium potential for chloride remained unchanged when the conductance response was depressed. The apparent or measurable rate constant of desensitization (KD) increased when the concentration of GABA increased, showing a saturation in KD-[GABA] relationship at higher concentration of GABA. The rate of desensitization did not change significantly when the resting membrane was hyperpolarized from -40 to -80 mV, though the conductance response was markedly depressed due to a characteristic voltage-dependence in the receptor activation (Matsumoto, 1982). These observations are discussed in terms of an hypothesis in which the desensitization of GABA receptor is the result of an additional binding of a GABA molecule to the activated receptor.