Prevention of chronic anthracycline cardiotoxicity in the adult Fischer 344 rat by dexrazoxane and effects on iron metabolism.

PURPOSE: Anthracyclines, such as doxorubicin and daunorubicin, continue to be widely used in the treatment of cancer, although they share the adverse effect of chronic, cumulative dose-related cardiotoxicity. The only approved treatment in prevention of anthracycline cardiotoxicity is dexrazoxane, a putative iron chelator. Previous in vitro studies have shown that disorders of iron metabolism, including altered IRP1-IRE binding, may be an important mechanism of anthracycline cardiotoxicity. METHODS: This study examined the role of IRP1-IRE binding ex vivo in a chronic model of daunorubicin cardiotoxicity in the Fischer 344 rat and whether dexrazoxane could prevent any daunorubicin-induced changes in IRP1 binding. Young adult (5-6 months) Fischer 344 rats received daunorubicin (2.5 mg/kg iv once per week for 6 weeks) with and without pretreatment with dexrazoxane (50 mg/kg ip). Other groups received saline (controls) or dexrazoxane alone. Rats were killed either 4 h or 2 weeks after the last dose of daunorubicin to assess IRP1-IRE binding. RESULTS: Contractility (dF/dt) of atrial tissue, obtained from rats 2 weeks after the last dose of daunorubicin, was significantly reduced in daunorubicin-treated compared to control rats. Dexrazoxane pretreatment protected against the daunorubicin-induced decrease in atrial dF/dt. However, left ventricular IRP1/IRE binding was not affected by daunorubicin treatment either 4 h or 2 weeks after the last dose of daunorubicin. CONCLUSIONS: IRP1 binding may not be altered in the rat model of chronic anthracycline cardiotoxicity.

Advanced integrated mouse YAC map including BAC framework.

Functional characterization of the mouse genome requires the availability of a comprehensive physical map to obtain molecular access to chromosomal regions of interest. Positional cloning remains a crucial way of linking phenotype with particular genes. A key step and frequent stumbling block in positional cloning is making a contig of a genetically defined candidate region. The most efficient first step is isolating YAC (Yeast Artificial Chromosome) clones. A robust, detailed YAC contig map is thus an important tool. Employing Interspersed Repetitive Sequence (IRS)-PCR genomics, we have generated an advanced second-generation YAC contig map of the mouse genome that doubles both the depth of clones and the density of markers available. In addition to the primarily YAC-based map, we located 1942 BAC (Bacterial Artificial Chromosome) clones. This allows us to present for the first time a dense framework of BACs spanning the genome of the mouse, which, for instance, can serve as a nucleus for genomic sequencing. Four large-insert mouse YAC libraries from three different strains are included in our data, and our analysis incorporates the data of Hunter et al. and Nusbaum et al. There is a total of 20,205 markers on the final map, 12,033 from our own data, and a total of 56,093 YACs, of which 44,401 are positive for more than one marker.

Sarcoplasmic reticulum calcium release is stimulated and inhibited by daunorubicin and daunorubicinol.

Cardiac effects of anthracyclines or their metabolites may include both the stimulation and inhibition of Ca(2+) release from sarcoplasmic reticulum. In this study, the ability of daunorubicin and its primary metabolite, daunorubicinol, to stimulate and inhibit Ca(2+) release from canine sarcoplasmic reticulum (SR) vesicles was investigated. It was observed that both daunorubicin and daunorubicinol were several fold more potent at inhibiting than they were at stimulating SR Ca(2+) release. Respective IC50 inhibition of daunorubicin and daunorubicinol for caffeine-induced calcium release was 1.2 and 0.6 microM, and for spontaneous Ca(2+) release was 3 and 1 microM. EC50's for daunorubicin- and daunorubicinol-induced calcium release were 30 and 15 microM, respectively. Inhibition of either spontaneous or caffeine-induced SR Ca(2+) release was inversely related to the amount of Ca(2+) loaded into the SR before exposure to daunorubicin or daunorubicinol. The free-radical scavenger dithiothreitol did not attenuate the ability of anthracyclines to inhibit SR Ca(2+) release. A nonquinone daunorubicin derivative, 5-iminodaunorubicin, was less potent than daunorubicin at inhibiting caffeine-induced Ca(2+) release. These data suggest anthracyclines and their metabolites may produce cardiotoxicity through free-radical independent, concentration-dependent effects on SR Ca(2+) release. These effects involve either inhibition or stimulation of SR Ca(2+) release and are partly dependent upon the presence of the quinone moiety.

Daunorubicin cardiotoxicity: evidence for the importance of the quinone moiety in a free-radical-independent mechanism.

Anthracyclines, such as daunorubicin (Daun), and other quinone-containing compounds can stimulate the formation of toxic free radicals. The present study tests the hypothesis that the quinone moiety of Daun, by increasing free-radical production, disrupts sarcoplasmic reticulum (SR) function and thereby inhibits myocardial contractility in vitro. We compared Daun with its quinone-deficient analogue, 5-iminodaunorubicin (5-ID), using experimental interventions to produce various contractile states that depend on SR function. At concentrations of Daun or 5-ID that did not alter contractility (dF/dt) of steady-state contractions (1 Hz) in electrically paced atria isolated from adult rabbits, only Daun significantly attenuated the positive inotropic effects on dF/dt of increased rest intervals (PRP; post-rest potentiation) or increased stimulation frequencies. Attenuation was to 98+/-6% at 1 Hz, and 73+/-8 and 67+/-8% for 30 and 60 sec PRP, respectively, and 73+/-3 and 63 +/-3% at 2 and 3 Hz, respectively, for 88 microM Daun (P<0.05, vs pre-drug baseline values, mean +/- SEM). These effects of Daun were similar to those of caffeine (2 mM), an agent well known to deplete cardiac SR calcium. We also examined the effect of Daun in isolated neonatal rabbit atria, which lack mature, functional SR; Daun did not alter the force-frequency relationship or PRP contractions. Additional studies in Ca(2+)-loaded SR microsomes indicated that both Daun and 5-ID opened Ca(2+) release channels, with Daun being 20-fold more potent than 5-ID in this respect. Neither anthracycline, however, induced free-radical formation in SR preparations (assayed via nicking of supercoiled DNA) prior to stimulating Ca(2+) release. Thus, our results indicate that Daun impairs myocardial contractility in vitro by selectively interfering with SR function; the quinone moiety of Daun appears to mediate this cardiotoxic effect, acting through a mechanism that does not involve free radicals.

Prevention by dexrazoxane of down-regulation of ryanodine receptor gene expression in anthracycline cardiomyopathy in the rat.

Anthracyclines can cause cumulative dose-related cardiotoxicity characterized by changes in Ca(2+) metabolism, including dysfunction of the sacroplasmic reticulum (SR) and decreased expression of Ca(2+)-handling proteins, such as the ryanodine receptor (RyR2). In this study, we examined the effect of dexrazoxane (ICRF-187), an iron chelator which prevents anthracycline cardiotoxicity, on RyR2 gene expression in rats treated chronically with daunorubicin. Daunorubicin (2.5 mg kg(-1) i.v. weekly for 6 weeks) produced cardiotoxicity as demonstrated by histopathologic changes. The ryanodine receptor/glyceraldehyde phosphate dehydrogenase (GAPDH) mRNA ratio was decreased by 38+/-3% (P<0.02) compared to values in control rats. Dexrazoxane pre-treatment (50 mg kg(-1); 1 h prior to each daunorubicin injection) prevented the decrease in RyR2/GAPDH mRNA ratio and histopathologic lesions in daunorubicin-treated rats. This is the first report that a protective agent such as dexrazoxane can ameliorate the decreased expression of a specific gene involved in anthracycline-induced cardiotoxicity.

A single amino acid of the human and rat neurotensin receptors (subtype 1) determining the pharmacological profile of a species-selective neurotensin agonist.

The neurotensin (NT) receptor, subtype 1 (NTR1), is a 7-transmembrane-spanning receptor, forming 3 extracellular and 3 intracellular loops. Previously, we showed that the third outer loop (E3) is the binding site for NT and its analogs, several of which bind with higher affinity to rat NTR1 (rNTR1) than to human NTR1 (hNTR1). In particular, NT34 [3,1'-naphthyl-l-Ala(11)]NT(8-13) has greater than 60-fold higher affinity for rNTR1 (46 and 60 pM for transiently- and stably-transfected cells, respectively) than for hNTR1 (2.8 and 5.8 nM for transiently- and stably-transfected cells, respectively) isolated from transfected cell membranes. Previously, our molecular modeling studies of rNTR1 and hNTR1 showed that the binding pocket in the human receptor for NT34 is smaller in volume from the bulky residue Tyr(339) in the pocket center, as compared with the corresponding residue Phe(344) in the rat binding pocket. Therefore, with site-directed mutagenesis, we derived mutant forms of rNTR1(F344Y) and hNTR1(Y339F). Examination of the mutant receptors from membranal preparations of transfected cells in radioligand binding assays and with intact cells in functional assays (phosphatidyl-4,5-bisphosphate turnover) showed that the human-like rat receptor and the rat-like human receptor bound NT34 with a predicted reverse of binding compared with its binding to the wild-type receptors. These results strongly affirm our molecular modeling studies and demonstrate the importance of the study of even minor structural variations in proteins to determine the basis of significantly different drug responses, an area of focus for pharmacological research in the 21st century.

A novel neurotensin peptide analog given extracranially decreases food intake and weight in rodents.

Neurotensin decreases food intake in the rat when injected into the cerebral ventricles. We tested the effect of a novel neurotensin analog (NT69L), injected intra-peritoneally (i.p.), on weight gain and food intake in rats. Sprague-Dawley rats (270 g) were injected i. p. with either saline or NT69L at 0.001 or 0.010 mg/kg. In further experiments, larger rats at a more steady state on the growth curve (400 g) were injected with either saline or 0.010 or 1 mg/kg NT69L. Food intake, water consumption and body weight were recorded daily. Weight gain was significantly reduced in the smaller rats injected with 0.001 or 0.010 mg/kg, showing only a 8.5 and 9.0% increase in original weight, respectively, as compared to a 29% increase for the controls. The larger rats injected with 1 mg/kg, had a significant reduction in body weight with a 3.0% decrease in original body weight as compared to a 2.4% increase for the controls. Food intake was significantly reduced suggesting that the weight loss observed after injection of NT69L was attributable in part to a reduction in food intake. The genetically obese Zucker rats injected with NT69L (1 mg/kg) had a significant reduction in weight gain and food intake. NT69L significantly increased blood glucose and corticosterone levels and decreased TSH and T4 in Sprague-Dawley and Zucker rats, an effect that was only transitory. NT69L also caused a decrease in norepinephrine in both the hypothalamus and nucleus accumbens, and an increase in dopamine, 3,4-dihydroxyphenylacetic acid (DOPAC) and serotonin. In this study, NT69L exhibited a consistent and dramatic effect on body weight and food intake in Sprague-Dawley and obese Zucker rats, and enabled us to study the role that NT plays in weight control and the functional interactions of NT with brain amines, and metabolic and endocrinological parameters.

Effects of a novel neurotensin peptide analog given extracranially on CNS behaviors mediated by apomorphine and haloperidol.

Neurotensin (NT) is a neuropeptide neurotransmitter in the central nervous system. It has been implicated in the therapeutic and in the adverse effects of neuroleptics. Activity of NT in brain can only be shown by direct injection of the peptide into that organ. However, we have developed a novel analog of NT(8-13), NT69L, which is active upon intraperitoneal (i.p.) injection. Like atypical neuroleptics, NT69L blocked the climbing behavior in rats, but not the licking and sniffing behaviors of a high dose (600 microgram/kg) of the non-selective dopamine agonist apomorphine. Its blockade of climbing was very potent with an ED(50) (effective dose at 50% of maximum) of 16 microgram/kg. Both apomorphine and NT69L caused a long-lasting hypothermia, which was greater with the peptide but not synergistic in combination with apomorphine. The ED(50) of NT69L for hypothermia was 390 microgram/kg. NT69L (up to 5 mg/kg i.p.) did not produce catalepsy. However, when given before haloperidol, NT69L, but not clozapine, completely prevented catalepsy. When given after haloperidol, NT69L, but not clozapine, reversed haloperidol's cataleptic effects with an ED(50) of 260 microg/kg. There was no significant difference between the ED(50)s for hypothermia and anticataleptic effects of NT69L. However, the ED(50) for blocking the effects of apomorphine was significantly lower than the other two. These data suggest that NT69L may have neuroleptic properties in humans and may be useful in the treatment of extrapyramidal side effects caused by typical neuroleptics such as haloperidol.

Analysis of binding sites and efficacy of a species-specific peptide at rat and human neurotensin receptors.

We have developed a neurotensin analog, L-[3,1'-naphthylalanine11]NT(8-13), NT34, that can distinguish between rat and human neurotensin receptors, and exhibits more than a 100-fold difference in binding affinities and a 60-fold difference in functional coupling to phosphatidylinositol turnover. Using cells transfected with different numbers of the appropriate receptors, we measured the changes in phosphatidylinositol production, and then evaluated the efficiency of receptor-effector coupling based on Furchgott's design. The binding of NT34 at both rat and human neurotensin receptors stably expressed in CHO-K1 cells was to two sites, while the binding of NT was to one site. At the rat receptor the equilibrium dissociation constant (Kd) for NT34 at the high-affinity site was 0.058 nM, while that at the low-affinity site was 3.1 nM. For the human receptor at the high-affinity site, the Kd for NT34 was 18 nM, while that at the low-affinity site was 180 nM. For both species the percentage of receptors representing the high-affinity site was approximately 60-70% with 30-40% at the low-affinity site. We derived agonist dissociation constants (Ka) for NT and NT34, which suggest that for NT34, the low-affinity site is functionally coupled to phosphatidylinositol turnover. Finally, we compared the relative efficacies of both compounds and found that NT34 was about 2-fold and 4-fold more efficacious than NT in stimulating phosphatidylinositol turnover in rat and human NT receptors, respectively.

Huprine X is a novel high-affinity inhibitor of acetylcholinesterase that is of interest for treatment of Alzheimer's disease.

Inhibitors of the enzyme acetylcholinesterase (AChE) slow and sometimes reverse the cognitive decline experienced by individuals with Alzheimer's disease. Huperzine A, a natural product used in traditional Chinese herbal medicine, and tacrine (Cognex) are among the potent AChE inhibitors used in this treatment, but the search for more selective inhibitors continues. We report herein the synthesis and characterization of (-)-12-amino-3-chloro-9-ethyl-6,7, 10,11-tetrahydro-7,11-methanocycloocta[b]quinoline hydrochloride (huprine X), a hybrid that combines the carbobicyclic substructure of huperzine A with the 4-aminoquinoline substructure of tacrine. Huprine X inhibited human AChE with an inhibition constant K(I) of 26 pM, indicating that it binds to this enzyme with one of the highest affinities yet reported. Under equivalent assay conditions, this affinity was 180 times that of huperzine A, 1200 times that of tacrine, and 40 times that of E2020 (donepezil, Aricept), the most selective AChE inhibitor currently approved for therapeutic use. The association and dissociation rate constants for huprine X with AChE were determined, and the location of its binding site on the enzyme was probed in competition studies with the peripheral site inhibitor propidium and the acylation site inhibitor edrophonium. Huprine X showed no detectable affinity for the edrophonium-AChE complex. In contrast, huprine X did form a ternary complex with propidium and AChE, although its affinity for the free enzyme was found to be 17 times its affinity for the propidium-AChE complex. These data indicated that huprine X binds to the enzyme acylation site in the active site gorge but interferes slightly with the binding of peripheral site ligands.

In vitro binding and CNS effects of novel neurotensin agonists that cross the blood-brain barrier.

Animal studies with neurotensin (NT) directly injected into brain suggest that it has pharmacological properties similar to those of antipsychotic drugs. Here, we present radioligand binding data for some novel hexapeptide analogs of NT(8-13) at the molecularly cloned rat and human neurotensin receptors (NTR-1), along with behavioral and physiological effects of several of these peptides after intraperitoneal (i.p.) administration in rats. One unique analog, NT66L, which had high affinity (0.85 nM) for the molecularly cloned rat neurotensin receptor (NTR-1), caused a drop in body temperature and antinociception at doses as low as 0.1 mg/kg after i.p. injection. At 30 min post-injection, the ED50 for NT66L-induced hypothermia (rectal temperature) and antinociception (hot plate test) was 0.5 and 0.07 mg/kg, respectively. At a dose of 1 mg/kg i.p., NT66L caused 100% of the maximum possible effect for antinociception for up to 2 h after administration. At this dose body temperature lowering was greater than -2.5 degrees C from 20 to 120 min after i.p. administration. These results in animals suggest that NT66L has agonist properties at NTR-1 in vivo after extracranial administration and provide support for its further study in behavioral tests predictive of neuroleptic activity.

Peptide nucleic acids targeted to the neurotensin receptor and administered i.p. cross the blood-brain barrier and specifically reduce gene expression.

Intraperitoneal injection of an unmodified antisense peptide nucleic acid (PNA) complementary to mRNA of the rat neurotensin (NT) receptor (NTR1) was demonstrated by a gel shift assay to be present in brain, thus indicating that the PNA had in fact crossed the blood-brain barrier. An i.p. injection of this antisense PNA specifically inhibited the hypothermic and antinociceptive activities of NT microinjected into brain. These results were associated with a reduction in binding sites for NT both in brain and the small intestine. Additionally, the sense-NTR1 PNA, targeted to DNA, microinjected directly into the brain specifically reduced mRNA levels by 50% and caused a loss of response to NT. To demonstrate the specificity of changes in behavioral, binding, and mRNA studies, animals treated with NTR1 PNA were tested for behavioral responses to morphine and their mu receptor levels were determined. Both were found to be unaffected in these NTR1 PNA-treated animals. The effects of both the antisense and sense PNAs were completely reversible. This work provides evidence that any antisense strategy targeted to brain proteins can work through i. p. delivery by crossing the normal blood-brain barrier. Equally important was that an antigene strategy, the sense PNA, was shown in vivo to be a potentially effective therapeutic treatment.

Refined radiation hybrid map of mouse chromosome 17.

We have made a radiation hybrid map of mouse Chromosome (Chr) 17 with 75 microsatellite markers, including those from McCarthy et al. (Genome Res 7, 1153-1161, 1997). Seventy-four of the markers are linked at LOD > 9, and all link at LOD > 5. A LOD 3 framework of 18 markers was used to construct a placement map. The order obtained is in good agreement with genetic maps, and distance estimates give an idea of how recombination rates vary across the chromosome. Recombination is remarkably low with respect to RH break frequency in the region from the centromere to the end of H2. This is similar in interspecific and intersubspecific crosses despite the inversion of a substantial part of this region in Mus spretus with respect to Mus musculus.

In vivo studies with low doses of levocabastine and diphenhydramine, but not pyrilamine, antagonize neurotensin-mediated antinociception.

The present study describes in vivo experiments in the rat addressing the role of levocabastine, and two other specific histamine H1 antagonists, diphenhydramine and pyrilamine, at neurotensin (NT)-mediated hypothermia and antinociception (hotplate). Levocabastine given i.p. or microinjected directly into the periaqueductal gray (PAG) did not cause antinociception or hypothermia. This indicates that despite the results with the recently-cloned levocabastine-sensitive NT receptors (NTR) in the rat (NTR-2) and mouse (NTRL), levocabastine by itself does not mediate either hypothermia or antinociception at NT receptors. However, pretreatment with 5 or 50 microg/kg of levocabastine or 5 microg/kg diphenhydramine all caused over a three-fold reduction in NT-mediated antinociception. Higher doses (500 or 5000 microg/kg) of levocabastine did not cause any antagonism of NT-mediated antinociception. All three antihistamines did not affect NT-mediated hypothermia. In addition, histamine H1 pathways are not involved in NT-mediated antinociception, as pretreatment with the much more potent histamine H1 antagonist pyrilamine did not affect antinociception mediated by NT. Therefore, these data may suggest the presence of yet unidentified NTR subtypes responsible for NT-mediated hypothermia and antinociception.

Evidence for additional neurotensin receptor subtypes: neurotensin analogs that distinguish between neurotensin-mediated hypothermia and antinociception.

Neurotensin (NT), a tridecapeptide, is a neurotransmitter that elicits potent effects including hypothermia and antinociception in mice and rats. To date, there are two types of the neurotensin receptor (NTR) that have been molecularly cloned from the rat. However, several lines of evidence suggest the presence of additional NTR subtypes. We have identified a NT analog of the NT(8-13) fragment, NT27, that selectively causes only the hypothermic response in vivo, when microinjected into the periaqueductal gray (PAG) of rats. A dose of 18 nmol of NT or NT27 caused a body temperature lowering of 1.8 and 1.2 degrees C, respectively. This same dose of NT or NT27 yielded a hotplate maximum physiological effect of 75% and 25%, respectively. Interestingly, despite its high KD (620 nM) at the cloned NTR-1, NT27-I (the iodinated form of NT27) exerted a potent hypothermic effect even at a very low dose (0.6 nmol). Equally intriguing, was that NT24, a sterioisomer of NT27, with a much higher affinity (KD=0. 5 nM) at NTR-1, did not selectively induce hypothermia in mice, but did selectively induce hypothermia in rats.

Specific gene blockade shows that peptide nucleic acids readily enter neuronal cells in vivo.

Peptide nucleic acids (PNAs) are DNA analogs that can hybridize to complementary sequences with high affinity and stability. Here, we report the first evidence of intracellular delivery of PNAs in vivo. Two CNS receptors, an opioid (mu) and a neurotensin (NTR-1), were targeted independently by repeated microinjection of PNAs into the periaqueductal gray. Behavioral responses to neurotensin (antinociception and hypothermia) and morphine (antinociception) were lost in a specific manner. Binding studies confirmed a large reduction in receptor sites. The loss of behavioral responses was long lasting but did fully recover. The implications of specifically and readily turning off gene expression in vivo are profound.

Aging and drug interactions. III. Individual and combined effects of cimetidine and cimetidine and ciprofloxacin on theophylline metabolism in healthy male and female nonsmokers.

The individual and combined effects of cimetidine and ciprofloxacin on theophylline metabolism were examined in healthy young and elderly male and female nonsmokers. Single-dose studies of theophylline pharmacokinetics were performed at base line and on the fifth day of each of three treatment regimens consisting of 400 mg cimetidine every 12 hr, 500 mg ciprofloxacin every 12 hr and the combination of cimetidine and ciprofloxacin. Base-line theophylline plasma clearance and formation clearance of theophylline metabolites decreased with age in both gender groups to a similar extent (20% less in elderly men than in young men; 24% less in elderly women than in young women). Individually, cimetidine and ciprofloxacin produced proportionate declines in plasma theophylline clearance that were similar among the four groups (range, 23.4-32.7% decrease). The combined regimen yielded further impairment in theophylline elimination compared with each agent alone (range, 35.9-42.6% decrease). Cimetidine was a nonselective inhibitor of theophylline metabolic pathways in young men, but it exerted a greater inhibitory effect on N-demethylation pathways in the other groups. Ciprofloxacin inhibited N-demethylations of theophylline to a greater extent than the hydroxylation pathway. Coadministration of these two inhibitors further reduced the formation of theophylline metabolites. The proportionate reduction in formation clearance of theophylline metabolites was similar among the four groups. Thus, the response to inhibition of theophylline metabolism by cimetidine and ciprofloxacin is not influenced by age or gender.

Age-related pharmacokinetics of daunorubicin and daunorubicinol following intravenous bolus daunorubicin administration in the rat.

Age-related differences in pharmacokinetics may be important in determining altered anthracycline cardiotoxicity in the senescent rat and also in older humans. This study examined the effect of aging on daunorubicin pharmacokinetics in the Fischer 344 rat. Daunorubicin 7.5 mg/kg was administered i.v. to 6- and 24-month-old male Fischer 344 rats and plasma and tissue sampling was performed over 168 h for assay of daunorubicin and daunorubicinol concentrations by high-performance liquid chromatography. Systemic clearance of daunorubicin was decreased in older compared to younger animals (56 +/- 4 versus 202 +/- 17 ml min-1 kg-1; P < 0.05). In addition, the area under the plasma daunorubicinol concentration/time curve was significantly increased in older rats. In the heart, the area under the concentration/time curve was significantly increased in senescence both in the case of daunorubicin (201 +/- 12 versus 86 +/- 4 micrograms h g-1; P < 0.05) and daunorubicinol (1347 +/- 118 versus 182 +/- 4 micrograms h g-1; P < 0.05). Furthermore, the peak mean concentrations of daunorubicin were increased in older compared to younger rats both in plasma (1078 +/- 82 versus 663 +/- 66 ng ml-1; P < 0.05) and in heart (27 +/- 1 versus 10 +/- 1 micrograms g-1; P < 0.05). This also was true for daunorubicinol in plasma (284 +/- 39 versus 168 +/- 27 ng ml-1; P < 0.05) and in myocardium (8.6 +/- 0.6 versus 2.4 +/- 0.2 micrograms g-1; P < 0.05). Following daunorubicin injection, the ratio of daunorubicinol to daunorubicin concentrations in tissues increased with time, particularly in plasma and heart in senescent rats. Thus, there are significant age-related changes in daunorubicin and daunorubicinol kinetics in the rat that could alter susceptibility to acute systemic toxicity and to chronic cardiotoxicity.