piperidines and 4-4--(2-(4-methylpiperidyl)-1-hydroxyethyl)biphenyl

4,4'-Bis-1-hydroxy-2-(4-methylpiperidin-1-yl)ethyl-biphenyl (A-4), a tertiary amine analog of hemicholinium-3 (HC-3), is an inhibitor of the sodium-dependent high-affinity choline uptake (HACU) system. We have evaluated 4-[1-hydroxy-2-(4-(18)F-fluoromethylpiperidin-1-yl)ethyl]-4'-[1-hydroxy-2-(4-methylpiperidin-1-yl)ethyl]biphenyl ((18)F-FA-4) and 4-[1-hydroxy-2-(4-(11)C-methylpiperazin-1-yl)ethyl]-4'-[1-hydroxy-2-(4-methylpiperidin-1-yl)ethyl]biphenyl ((11)C-pipzA-4), an (18)F- and a (11)C-labeled derivative of A-4 as potential in vivo tracers for the HACU system.. The biodistribution of both compounds was determined in mice, and the intracerebral distribution was visualized by ex vivo and in vitro autoradiography. The in vitro affinity of the compounds was determined by a displacement study with (3)H-HC-3 on mice brain slices.. In mice, both tracers show a high and persistent brain uptake. In vitro autoradiography shows binding to the striatum, whereas ex vivo autoradiography shows homogeneous binding throughout the brain. FA-4 and pipzA-4 inhibited the (3)H-HC-3 binding with a 50% inhibitory concentration of 57 nmol/L and 320 nmol/L, respectively.. The evaluated compounds have affinity for HACU and show high uptake in the brain. In vitro binding of the compounds to the striatum cannot be inhibited by the presence of HC-3, whereas binding of HC-3 was inhibited by the presence of both FA-4 and pipzA-4, suggesting allosteric binding.

Topics: Animals; Autoradiography; Biphenyl Compounds; Carbon Radioisotopes; Cerebellum; Corpus Callosum; Corpus Striatum; Deuterium; Fluorine Radioisotopes; Hemicholinium 3; Kidney; Liver; Lung; Male; Membrane Transport Proteins; Mice; Myocardium; Piperidines; Radioligand Assay; Radiopharmaceuticals; Telencephalon; Tissue Distribution

A-4, A-5 and HC-3 are experimental bis tertiary and quaternary amines which have been shown to be potent inhibitors of the sodium-dependent, high affinity choline uptake system. When incubated with neuroblastoma cells, experimental compounds A-4, A-5 and HC-3 inhibit choline metabolism. Over a 24-hr incubation, A-4, A-5 and HC-3 produced a significant decrease in total choline accumulation, choline incorporation into phospholipid and free choline content. However, despite decreases in choline incorporation into phospholipid, no change occurred in content of phosphatidylcholine. Treatment of cells with A-4, A-5 and HC-3 resulted in an increase in the incorporation of S-adenosyl-methionine into phosphatidylcholine. However, the incorporation of ethanolamine or serine into phosphatidylcholine was not increased. Phosphatidylcholine turnover was decreased in cells treated with A-4 and A-5. A-4, A-5 and HC-3 produce significant decreases in choline metabolism; however, the cells are able to maintain membrane integrity by decreasing turnover of phosphatidylcholine and increasing phosphatidylcholine synthesis through the methylation pathway. These studies suggest that the biological effects of A-4 and A-5 are independent of membrane perturbations.

Topics: Animals; Biphenyl Compounds; Choline; Ethanolamine; Ethanolamines; Mice; Neuroblastoma; Phosphatidylcholines; Phospholipids; Piperidines; S-Adenosylmethionine; Tumor Cells, Cultured

The neuroblastoma-glioma hybrid cell (NG108-15) has a sodium-dependent, high-affinity choline transport system with a Km of 16.0 +/- 3.4 microM and a Vmax of 214.5 +/- 27.7 pmol/min/mg protein. A-4, A-5 and HC-3 produce dose-dependent inhibition of high-affinity choline transport in NG108-15 cells. Following 24 h exposure to approximately the EC50 of each inhibitor, no significant decrease was found in total choline accumulation or in choline incorporation into phosphatidylcholine. However, when additional inhibitor was added during the 24 h incubation, significant decreases in choline accumulation were produced by A-4 and A-5. Following 24 h exposure to each compound, only A-4 was able to significantly affect free choline content. In contrast, each inhibitor was able to significantly decrease acetylcholine content following 24 h exposure. Possible reasons for consistent decreases in acetylcholine versus minimal changes in choline metabolism will be discussed.

Topics: Acetylcholine; Animals; Biological Transport, Active; Biphenyl Compounds; Choline; Hemicholinium 3; Neurons; Parasympatholytics; Piperidines; Tumor Cells, Cultured

A-4 and A-5 are tertiary and N-methyl quaternary 4-methylpiperidine analogs of hemicholinium-3 (HC-3). Previous work in this laboratory has shown A-4 and A-5 to be inhibitors of the sodium-dependent, high affinity choline uptake system (SDHACU). Their effects on choline transport were characterized further using neuroblastoma 41A3 cells. These cells rapidly take up choline through two separate mechanisms: a SDHACU system and a sodium-independent, low affinity uptake system (SILACU). A-4, A-5 and HC-3 decreased 5 microM choline transport in a dose-dependent fashion. The compounds were unable to decrease choline transport at 250 microM choline suggesting that they are inactive with respect to SILACU. All three compounds significantly increased the Km but not the Vmax for the SDHACU system, suggesting a competitive mechanism of inhibition. Ki values ranged from 18 to 25 microM for A-4, 20 to 26 microM for A-5 and 68 to 75 microM for HC-3. Dose-response curves for inhibition of choline transport by A-5 and HC-3 were not changed by a 24-hr pre-exposure of the cells to each inhibitor. However, after a 24-hr pre-exposure to A-4, a significantly different dose-response curve was obtained compared to the dose-response curve for A-4 in untreated cells. After a 24-hr pre-exposure, a 4-hr recovery period was sufficient to remove the effect of each compound. These data suggest that A-4 and A-5, like HC-3, inhibit the SDHACU, competitively and reversibly.

Topics: Animals; Biological Transport; Biphenyl Compounds; Choline; Dose-Response Relationship, Drug; Mice; Neuroblastoma; Parasympatholytics; Piperidines; Sodium; Tumor Cells, Cultured

Rats were injected with either saline; A-4 (40 mg/kg, i.p.), a bis tertiary amine derivative of hemicholinium-3; or A-5 (50 micrograms/kg, i.p.), a bis quaternary amine derivative of hemicholinium-3, 1 h prior to moderate fluid percussion brain injury. A variety of reflexes and responses were measured up to 60 min following injury, and body weight and several neurological measures were taken daily up to 10 days following injury. Pretreatment with either A-4 or A-5 significantly attenuated components of transient behavioral suppression, as well as more enduring deficits in body weight and beam walk and beam balance performance. A-4 administered prior to fluid percussion was found to reduce striatal, but not pontine, acetylcholine content. A-5 did not significantly reduce acetylcholine content in either area. Both A-4 and A-5 pretreatment prevented a significant increase in acetylcholine content in the cerebrospinal fluid following fluid percussion injury; however, only A-5 significantly reduced plasma acetylcholine content. These results confirm cholinergic involvement in the production of both transient and longer-lasting behavioral deficits following traumatic brain injury. Furthermore, traumatic brain injury may allow plasma constituents to gain access to the central nervous system.

Topics: Acetylcholine; Animals; Biphenyl Compounds; Brain Injuries; Cholinergic Fibers; Male; Movement Disorders; Piperidines; Rats; Rats, Inbred Strains

The in vitro effects of a tertiary amine, 4-methyl piperidine analog of hemicholinium-3 (A-4), were investigated on acetylcholine (ACh) metabolism in rat striatal slices. Rat striatal slices were incubated in the presence of 1.0 microM [3H]-choline in the presence or absence of 0.1 mM A-4. High pressure liquid chromatography with electrochemical detection was utilized to separate and measure total and [3H]-labeled ACh and choline. The effects of A-4 on [3H]-choline uptake, ACh and choline content, ACh release, and specific activity of ACh and choline tissue pools were investigated. Results indicated that A-4 inhibited the uptake of [3H]-choline into the striatal slices. Addition of 0.1 mM A-4 also produced a significant reduction in ACh content and ACh release and reduced the specific activity of the tissue choline and ACh pools. The effects of A-4 were more prominent upon immediate incubation than after a 60-min preincubation. These studies demonstrate that the pharmacological effects exhibited by A-4 are consistent with inhibition of choline uptake, with subsequent reduction in ACh synthesis and release.

Topics: Acetylcholine; Animals; Biphenyl Compounds; Choline; Chromatography, High Pressure Liquid; Corpus Striatum; Hemicholinium 3; Piperidines; Potassium; Rats; Structure-Activity Relationship

Previous studies have shown that quaternary and tertiary 4-methyl piperidine derivatives of hemicholinium-3 (A-5 and A-4, respectively) are potent inhibitors of choline uptake. The d-, l-, and mesostereoisomers of A-5 and A-4 were separated and the potency and reversibility were compared. Isomeric forms of each compound were found to be approximately equipotent inhibitors in the following preparations: inhibition of rabbit neuromuscular transmission using the sciatic nerve-gastrocnemius muscle preparation, reductions in acetylcholine content in rat caudate tissue slices and inhibition of choline uptake in neuroblastoma cells, line NB41A3. Because these results show no difference in potency or reversibility for the stereoisomers of A-5 or A-4, these studies indicate that hydroxyl substitutions in these agents do not play a role in their biologic activity. Perhaps only 2-point attachment is required for inhibition of choline transport by hemicholinium-like compounds.

Topics: Acetylcholine; Animals; Biphenyl Compounds; Caudate Nucleus; Cells, Cultured; Choline; Hemicholinium 3; Male; Piperidines; Rabbits; Rats; Rats, Inbred Strains; Sciatic Nerve; Stereoisomerism