A study of the behaviour of cobalt chloride during the labelling of leukocytes with 99Tcm-HMPAO stabilised in vitro by the addition of cobalt chloride solution.

The method described by Weisner et al. for stabilizing 99Tcm-hexamethylpropylene amine oxime (99Tcm-HMPAO) with cobalt chloride hexahydrate solution is the most promising developed to date. The aim of our work was to study the behaviour of cobalt during the labelling of leukocytes with 99Tcm-HMPAO stabilized in vitro using this method. Three parallel labellings were carried out using six blood samples taken from polycythaemic patients. The first set of labellings was performed using 99Tcm-HMPAO, the second set using 99Tcm-HMPAO stabilized with cobalt chloride hexahydrate solution "spiked' with 57Co-chloride, and the third set using 57Co-chloride solution alone. Measurements of radioactivity content were made on the leukocyte pellets after washing at five time points post-labelling (t = 0, 30 min, 1 h, 2 h and 4 h). The data show that leukocytes do not retain cobalt during labelling with stabilized 99Tcm-HMPAO. Studies carried out in parallel demonstrated that the presence of cobalt in the cell-labelling medium had no effect on cell viability.

Mechanism of uptake of technetium-tetrofosmin. I: Uptake into isolated adult rat ventricular myocytes and subcellular localization.

BACKGROUND: 99mTc-labeled tetrofosmin is a new myocardial imaging agent that gives stable heart uptake. However, little is known about the mechanism of uptake in heart tissue. METHODS AND RESULTS: Uptake of 99mTc-labeled tetrofosmin has been examined in isolated adult rat ventricular myocytes. The time course of uptake, efflux rate, and the effect of metabolic and cation channel inhibitors have been assessed. The subcellular localization of radioactivity in ex vivo rat heart tissue was examined by differential centrifugation of ventricular homogenate. Uptake into rat myocytes was found to be rapid and plateaued at approximately 1.5 pmol/10(6) cells/nmol extracellular Tc-labeled tetrofosmin after 60 minutes of incubation. Uptake was temperature dependent but independent of extracellular Tc-labeled tetrofosmin concentration. Uptake at 30 minutes was inhibited by the metabolic inhibitors iodoacetic acid acid and 2,4-dinitrophenol protein but was not affected by cation channel inhibitors. Cells previously incubated with 99mTc-labeled tetrofosmin and then placed into fresh medium were found to have a slow efflux of activity; after 1 hour, 65% of activity was still cell associated. The localization of radioactivity in subcellular fractions indicated that the majority of activity was recovered with the cytosol. However, examination of the distribution of two mitochondrial enzymes indicated that this may have been artifactual. Use of carbonyl cyanide m-chlorophenyl-hydrazone or oligomycin to perturb mitochondrial membrane potential decreased or increased recovery in the mitochondrial fraction, respectively. CONCLUSIONS: 99mTc-labeled tetrofosmin uptake by myocytes is by a metabolism-dependent process that does not involve cation channel transport. The most likely mechanism for this is by potential driven diffusion of the lipophilic cation across the sarcolemmal and mitochondrial membranes.

The radiation dosimetry of 99Tcm-exametazime.

The biodistribution of the regional cerebral perfusion imaging agent, 99Tcm-exametazime, has been studied with volunteer subjects at Aberdeen, Homburg, Manchester and Milan. Data from these studies have been pooled and analysed to formulate a kinetic biodistribution model, allowing estimation of time integrals of activity in various body organs. Estimates of radiation dose to humans injected with this material have been made by applying the MIRD formalism to these data. The highest doses occur to the lachrymal glands, gallbladder and kidneys (33, 27 and 18 mSv, respectively, per 500 MBq administered.) The lachrymal glands were visualized in only 6 of the 26 volunteer studies. The effective dose equivalent, for the worst case individual, is 8.3 mSv. In the majority of subjects where there was no uptake in the lachrymal gland, the effective dose equivalent reduces to 6.9 mSv.

Technetium-99m d,l-HM-PAO: a new radiopharmaceutical for SPECT imaging of regional cerebral blood perfusion.

Following investigation of a large number of new ligands based upon propylene amine oxime (PnAO) the d,l-diastereoisomer of hexamethyl propyleneamine oxime (HM-PAO) was selected as the preferred ligand for 99mTc as a tracer for cerebral perfusion imaging. The neutral, lipophilic 99mTc complex of d,l-HM-PAO was formed in high yield by stannous reduction of 99Mo/99mTc generator eluate using a kit formulation of the ligand. Two minutes following i.v. administration of this complex in rats, 2.25% of the injected dose appears in the brain. Little washout of the tracer is observed up to 24 hr postinjection. By qualitative autoradiographic comparison with iodoantipyrine this new radiopharmaceutical displays blood flow dependent brain uptake with little redistribution of the tracer over time. The lipophilic 99mTc complex converts slowly in vitro to a secondary complex. This conversion process may account for the ability of [99mTc]d,l-HM-PAO to be retained within the brain without redistribution.

A comparative study of the brain uptake and early kinetics of 99mTc-dl HM-PAO and other PnAO derivatives in baboons.

Derivatives of propylene-amine-oxime (PnAO) have been synthesized which form a neutral lipid-soluble complex with 96mTc and can be supplied as freeze-dried kits. The complexes cross the intact blood-brain barrier. This report shows the brain uptake, early kinetics and biodistribution in normal adult baboons of 5 99mTc-PnAO derivatives and 2 isomers of one of the tested derivatives (HM-PAO). The brain uptake of the favoured dl-isomer of HM-PAO reaches its maximum of 4.3% (whole brain/whole body) 1 min p.i. and a clearance of less than 8% was observed 23 min p.i.

Technetium-99m HM-PAO stereoisomers as potential agents for imaging regional cerebral blood flow: human volunteer studies.

A total of nine normal volunteer subjects were studied with three forms of [99mTc] hexamethylpropyleneamineoxime (HM-PAO), a potential cerebral blood flow imaging agent. One, the d,l isomer, showed 4.1% uptake in the brain which remained constant over 8 hr. There was good differentiation between uptake in gray and white matter on tomographic slices. We propose that this agent may allow regional cerebral blood flow imaging to be performed on a routine basis.

Development of a 99Tcm-labelled radiopharmaceutical for cerebral blood flow imaging.

A new radiopharmaceutical, 99Tcm-hexamethylpropyleneamine oxime (99Tcm-HM-PAO) has shown considerable promise for single photon emission tomographic (SPECT) imaging of the brain. In animal biodistribution studies this complex demonstrates good brain uptake with prolonged retention of activity in brain. Further improvement of these properties, resulting in higher brain uptake with very slow washout and fixed regional distribution has been achieved following the isolation of the d,1-diastereoisomer of HM-PAO.

A 99Tcm-labelled radiotracer for the investigation of cerebral vascular disease.

The first clinical data is given on 99Tcm-hexamethylpropyleneamine oxime (HM-PAO) in normal subjects and patients with established stroke. Regional cerebral blood flow maps (rCBF) have been recorded and displayed in tomographic mode with this new radiopharmaceutical. Good images were obtained, comparable to those achieved with 123I-isopropylamphetamine (IMP).

Technetium thiodiglycollic acid (99mTc-TDG). A new radiopharmaceutical with the potential for the assessment of renal function.

A mixture of hydrophilic complexes is formed on the reduction (employing tin metal as a reductant) of sodium pertechnetate 99mTc in the presence of the ligand thiodiglycollic acid (TDG). When administered to rats, the mixture demonstrated a renal clearance rate marginally greater than Glomerular filtration rate (GFR). HPLC analysis indicated the formation of two technetium complexes of TDG. After isolation of the complexes and their administration to rats, one (complex 1) showed renal clearance similar to that of 99mTc Diethylenetriamine pentaacetic acid (DTPA), while the other (complex 2) demonstrated renal clearance similar to that of 125I o-iodo hippuric acid. On heating the mixture of complexes, the proportion of the faster-clearing 99mTc TDG complex increased to 92% of the total activity, and the biodistribution of the material following heat rearrangement was equivalent to that of isolated complex 2.

Studies on the persistence of basic amines in the rabbit lung.

We have investigated the time-course of the pulmonary deposition of imipramine (IMIP) following a single iv injection into rabbits. A pool of IMIP and its demethylated metabolites, which exhibited considerable persistence (half-life of decay = 4 hr), was formed in the lung. This pool, now called the slowly effluxable pool (SEP), appears to be synonymous with the noneffluxable pool (NEP), which we have previously shown to be formed with IMIP in the isolated perfused lung (PL). Furthermore, this pool appears to be responsible for the pulmonary persistence of IMIP, inasmuch as 24 hr after an iv injection it contributes greater than 90% of the total lung concentration. Chlorphentermine and methadone formed SEP's in the IPL of comparable size to that formed with IMIP, whereas phentermine formed a considerably smaller SEP. These results suggest that the degree of hydrophobicity of the amine is an important determinant of the size of the SEP formed. This further supported by the lack of an SEP with the relatively polar amines, 5-hydroxytryptamine and amphetamine. The 10-fold difference in the size of the SEP for compounds known to induce pulmonary phospholipidosis (chlorphentermine and IMIP) and known not to induce lipidosis (phentermine and amphetamine) may suggest a possible involvement of the SEP in the onset of phospholipidosis; this possibility is discussed.

The pharmacodynamics of 5-hydroxytryptamine uptake and metabolism by the isolated perfused rabbit lung.

We have investigated the dynamics for the removal of 5-hydroxytryptamine (5-HT) from the circulation using the isolated perfused rabbit lung. 5-HT was removed from the circulation at a constant rate and metabolized completely to 5-hydroxyindoleacetic acid which effluxed from the lung into the circulation. Two methods were developed to determine the constant rate of removal of 5-HT: (1) the constant rate is equal to the difference between the 5-HT concentration flowing into the lung and the 5-HT concentration in the effluent times the flow rate and (2) extrapolation of the rate of appearance of radioactivity in the effluent to zero time. With these methods, we have confirmed the 5-HT is removed by the lung by a carrier-mediated Na+-dependent transport system. Studies of transport systems in perfused organs required an adequate supply of the chemical to the lung. Supply rates less than removal rate will result in erroneous measurements of the constant removal rate. The relationships between the rate of removal, perfusate concentration and perfusion rate were analyzed.

A study of the dynamics of imipramine accumulation in the isolated perfused rabbit lung.

An isolated perfused rabbit lung preparation (IPL) was used to study the uptake, accumulation, and efflux of the tricyclic amine imipramine (IMIP). The rate of IMIP uptake into the IPL was resolved into two exponential components (rates 1 and 2 of uptake). The initial velocities for these uptake components were linearly related to the concentration of IMIP in the perfusate (Cp). This linear relationship indicates that IMIP accumulated in the IPL by diffusion and/or binding. The steady-state accumulation of IMIP was obtained by integration of the exponential expression relating the rate of IMIP uptake to time. The amount of IMIP accumulated at steady state by rate 1 was linearly related to Cp, whereas the amount of IMIP accumulated at steady state by rate 2 was saturable with respect to Cp. These steady-state data are in agreement with the steady-state accumulation data previously reported from experiments with the recirculating blood-perfused rabbit lung. In the absence of IMIP in the perfusate. IMIP that had previously accumulated in the IPL effluxed from the lung at three rates (t1/2 = 18 sec, 58 sec, and 8.25 min), which indicates that accumulated IMIP was in at least three pools in the lung. In addition, a noneffluxable pool was detected which was not the result of irreversible binding to tissue.ated in pools 1 and 2 by rate 1 of uptake. The IMIP in pool 3 and in the noneffluxable pool was accumulated in the IPL by rate 2 of uptake. Efflux pool 3 and the IMIP accumulated at a steady state by rate 2 of uptake were resolved by a Hofstee plot into a biphasic curve indicating two types of binding sites. The noneffluxable pool of IMIP in the IPL was saturable with increasing Cp and represented approximately 30% of the IMIP accumulated by rate 2 into pool 3. Rate 2 of uptake, pool 3, and particularly the noneffluxable pool of IMIP in the IPL are possibly responsible for the accumulation and persistence of Imip in the lung as seen in whole body distribution studies. The physicochemical properties of IMIP and other compounds known to be accumulated in lung tissue are discussed in relation to the possible involvement of lung phospholipids and the compartmentalization of IMIP in the concentric lamellar organelles of the lung.