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In order to examine the relationship between long-term potentiation (LTP) and phosphoinositide (PI) turnover, we evaluated these throughout anesthetized rat brain using carbon-11-labeled diacylglycerol (11C-DAG). High-frequency tetanic stimulation (400 pulses at 400 Hz) to the perforant pathway induced LTP in rat dentate gyrus. In autoradiograms of rat brains, LTP was associated with the occurrence of multiple highly radioactive spots in many regions distant from the stimulated site. Following i.v. administration of an NMDA receptor antagonist prior to stimulation, however, no high-density spots were found. These findings directly demonstrate that potentiation of phosphoinositide-derived signaling was induced during LTP, and the finding of multiple location suggests the occurrence of polysynaptic neurotransmission through neural networks pertaining to learning and memory.

Phospholipid metabolism is closely related to membrane perturbation in cerebral ischaemia. We investigated in vivo topographical lipid metabolism using an iodine-123-labelled diacylglycerol analogue, (1-(15-(4-iodine-123-iodophenyl)-pentadecanoyl)-2-stearoyl-rac-gly cerol) (123I-labelled DAG), in a middle cerebral artery (MCA) occlusion model with the aim of positive imaging of ischaemic insult. Sprague-Dawley rats underwent coagulation of the MCA to induce permanent occlusion. MCA occlusion times prior to injection of 123I-labelled DAG ranged from 15 min to 14 days. Each rat was injected with 11-37 MBq of 123I-labelled DAG via a tail vein. After 30 min, in vivo autoradiographs were reconstructed. Scanning of the living rat brain in this MCA occlusion model was performed using a gamma camera with a pinhole collimator. Cerebral infarctions were recognized in the frontal cortex, the parietal cortex and the lateral portion of the caudate-putamen by 2,3,5-triphenyltetrazolium hydrochloride staining. In infarcted regions (region 1), 123I-labelled DAG incorporation showed a slight decrease up to 12 h; it then increased up to 6 days and decreased thereafter. In peri-infarcted regions (region 2), the incorporation showed almost no change up to 12 h, then increased up to 5-6 days and decreased thereafter. In other regions (region 3), the incorporation showed no change. Lipid analysis showed that 123I-labelled DAG was metabolized to 15-(4-iodine-123-iodophenyl)-pentadecanoic acid by DAG lipase and to 123I-labelled phosphatidylcholine. Scanning of the ischaemic region showed higher accumulation than on the non-lesioned side. We established a method to visualize ischaemic foci as positive images. The early changes in 123I-labelled DAG incorporation were closely related to DAG lipase, which degraded the accumulated intrinsic DAG, and increased 123I-labelled DAG incorporation in the chronic stage involves several aspects of neural destruction in the process of autolysis. It is concluded that the reported method could have a clinical future.

Specific hemodynamic changes in acute ischemia were investigated using a middle cerebral artery occlusion primate model and positron emission tomography. The cerebral blood flow (CBF), cerebral blood volume, oxygen extraction fraction (OEF), and cerebral metabolic rate for oxygen were measured 1, 3, and 9 hours after occlusion. OEF showed an increase in ischemic areas, and especially where CBF was below 18 ml/100 gm/min 1 hour after occlusion the OEF increased significantly (0.69 +/- 0.20, p < 0.05). Nine hours after occlusion, the OEF values were lower compared to those 1 and 3 hours after occlusion. Areas where CBF ranged from 18 to 31 ml/100 gm/min showed an increase in OEF at all times (p < 0.05). Clearly, OEF changes remarkably in the acute stage.

A new synthesis of (C-11)propyl ketene using a HCl/Helium gas mixture and glass beads was developed. This improved synthesis method gave high yields and good reproducibility, which had been difficult to attain with our previous method. The (C-11) propyl ketene is a useful precursor for labeling, in high specific activity, bioactive compounds which have a hydroxyl group. The reactivity of (C-11) propyl ketene in the acylation of various alcohols was evaluated. The relative reactivity of primary, secondary and tertiary alcohols were 1, 0.4 and 0.1, respectively, and the reactivity of the noncarrier added conditions was similar to that using cold propyl ketene. This (C-11)propyl ketene was also useful for the formation of N-butyl compounds, making it possible to label many bioactive compounds.

Positron emission tomography (PET) radiotracers have very short physical half-lives. It is hard to complete a bacterial endotoxins test prior to release from medical institutes. For endotoxin quantitative determination, limulus amebocyte lysate (LAL) reagent and kinetic-turbidimetry system were previously developed. We investigated the possibility of a short time test by means of positively charged filters. As a result of this study, the effects of positively charged filters on endotoin removal were over 99.5% for [18F]FDG and [18F]NaF, which were contaminated with the indicated concentration of endotoxin. Combining this filter and the kinetic-turbidimetric method, it was possible to complete a bacterial endotoxins test in 5 min prior to the patient's administration. This test should be required prior to release for PET radiopharmaceutical quality control. It has been suggested that this combination is a good method for this purpose.

We recently reported that myocardial phosphoinositide (PI) metabolism can be visualised by 1-[1-11C]-butyryl-2-palmitoyl-rac-glycerol (11C-DAG) in rats with myocardial infarction (MI). Angiotensin II, the receptors for which are expressed predominantly in infarcted areas with active fibrogenesis rather than in non-infarcted regions, is involved in the upstream signalling systems of PI metabolism and plays an important role in the process of left ventricular (LV) remodelling after MI. We therefore hypothesised that the distribution of 11C-DAG after MI may be affected by the inhibition of angiotensin converting enzyme, which is one of the most important factors in the development of LV remodelling after MI. Rats were injected with 11C-DAG after 3 or 10weeks of treatment with captopril or no treatment following coronary artery ligation, and quantitative autoradiography was performed. Cells occupying the infarcted region were identified by immunohistochemistry. Compared with untreated rats, treatment with captopril for 3 weeks after MI elicited a reduction in the 11C-DAG uptake in the infarcted region (P

Phosphoinositide turnover is closely connected to modulation of synaptic function and is part of an important second messenger-producing system. New radioligands for imaging second messenger systems by positron emission tomography have been developed: carbon-11-labeled 1,2-diacylglycerols. The theoretical background of second messenger imaging is described in detail and the relation between the biologically active compounds and potential tracers for imaging second messenger systems is discussed. We report informative findings on postsynaptic biological responses in the living human brain of healthy normal subjects and with various diseases.

ANTISENSE phosphorothioate oligodeoxynucleotide for mRNA of glial fibrillary acidic protein (GFAP) was labeled with the positron emitter 1 C and administered i.v. to rats bearing glioma, which were expected to exhibit active expression of GFAP. Antisense oligodeoxynucleotide was retained in tumor cells, yielding clear images of tumors, while the control 20% mismatch oligodeoxynucleotide and sense-strand oligodeoxynucleotide were not retained in tumor cells. Findings revealed sequence-specific binding of the antisense oligodeoxynuc eotide to the GFAP mRNA. Our methods can be used directly for non-invasive imaging of human gene expression using PET, a frequently used method of clinical examination.

ALZHEIMER'S disease (AD) is a region-specific degenerative disease that mainly impairs the temporal and parietal lobes, with preservation of the frontal lobes until advanced stages of disease. Since [ 11 C]diacylglycerol PET facilitates examination of loci exhibiting plasticity, it was performed in eight patients with AD and six age-matched normal control subjects to evaluate frontal lobe function. [ 11 C]Diacylglycerol tomograms obtained from patients with AD demonstrated strong spotty incorporation of [ 11 C]diacylglycerol mainly in the frontal association areas. However, [ 18 F]fluorodeoxyglucose tomograms exhibited region-specific findings such as decreased CMRGlc in the parietotemporal association areas, which are involved in impairment of cognitive function. The strong spotty incorporation of [ 11 C]diacylglycerol suggested a compensatory plastic process in the frontal lobes in response to involvement by Alzheimer's disease of the posterior association areas.

A new method was developed to synthesize positron labeled phorbol esters for second messenger imaging based on a receptor binding to protein kinase C. [1-11C]propyl ketene was produced by the pyrolytic decomposition of [1-11C]butyric acid. This new method was made possible by the reaction of [1-11C]propyl ketene to the hydroxyl group of phorbol under the non-carrier added conditions. The precursor protected in the 20-hydroxyl group of phorbol was acylated by [1-11C]propyl ketene to the 13-hydroxyl group producing phorbol 13-[11C]butyrate ([11C]P13Bu). Total synthesis time was about 60 minutes and the specific activity was greater than 67 GBq/μmol (1800 mCi/μmol). [11C] P13Bu will prove to be useful ligand for PET studies due to the simple procedures required for synthesis.

Chronological changes in regional cerebral blood flow (rCBF) and in the regional cerebral metabolic rate for glucose (rCMRGIu) were studied by a double tracer autoradiographic method in regions of local ischemia in rabbit brains. Local ischemia was produced by cautery of the bilateral vertebral artery, followed several days later by cautery of the left middle cerebral artery through a transorbital approach and ligation of the left common carotid artery. Autoradiography was performed, 2 hours, 6 hours, and 4 days after occlusion, by a double tracer method involving the use of 14Ciodo-antipyrine and 18F-fluoro-deoxyglucose. Absolute rCBF values were estimated by Sakurada's method and rCMRGIu values by Hutchins' formula. Histological examination was performed concurrently with the rCBF and rCMRGIu study. Mildly ischemic lesions (rCBF of 25 to 40 ml/100 g/ min) were detected in the superior portion of the left frontal lobe, the left parietal lobe, and the left occipital lobe. Severely ischemic lesions (rCBF below 25 ml/100 g/min) were found in the lateral part of the left frontal lobe, the left temporal lobe, and the left caudate nucleus. In the mildly ischemic regions, rCMRGIu decreased in proportion to the decrease in rCBF. That is, matched low perfusion was observed, but there were no histological abnormalities. In severe ischemia the situation was quite different. Two hours after occlusion, most areas showed a decrease in rCMRGIu in proportion to the decrease in rCBF. However, 6 hours after occlusion, rCMRGIu decreased nonuniformly: in some places the decrease was dramatic and in others, rCMRGIu residue was found. Four days after occlusion, the reduction in rCMRGIu was again proportional to the rCBF decrease. In severely ischemic regions, necrosis was observed 4 days after occlusion. Thus, disturbance of glucose metabolism and eventual tissue necrosis occurred in severely ischemic regions. Moreover, the results suggest that nonuniformity of the decrease in rCMRGIu may have prognostic significance in cases of severe ischemic brain insult.