Perinatal Fentanyl Exposure Leads to Long-Lasting Impairments in Somatosensory Circuit Function and Behavior.

One consequence of the opioid epidemic are lasting neurodevelopmental sequelae afflicting adolescents exposed to opioids in the womb. A translationally relevant and developmentally accurate preclinical model is needed to understand the behavioral, circuit, network, and molecular abnormalities resulting from this exposure. By employing a novel preclinical model of perinatal fentanyl exposure, our data reveal that fentanyl has several dose-dependent, developmental consequences to somatosensory function and behavior. Newborn male and female mice exhibit signs of withdrawal and sensory-related deficits that extend at least to adolescence. As fentanyl exposure does not affect dams' health or maternal behavior, these effects result from the direct actions of perinatal fentanyl on the pups' developing brain. At adolescence, exposed mice exhibit reduced adaptation to sensory stimuli, and a corresponding impairment in primary somatosensory (S1) function. In vitro electrophysiology demonstrates a long-lasting reduction in S1 synaptic excitation, evidenced by decreases in release probability, NMDA receptor-mediated postsynaptic currents, and frequency of miniature excitatory postsynaptic currents (mEPSCs), as well as increased frequency of miniature inhibitory postsynaptic currents (mIPSCs). In contrast, anterior cingulate cortical neurons exhibit an opposite phenotype, with increased synaptic excitation. Consistent with these changes, electrocorticograms (ECoGs) reveal suppressed ketamine-evoked γ oscillations. Morphologic analysis of S1 pyramidal neurons indicate reduced dendritic complexity, dendritic length, and soma size. Further, exposed mice exhibited abnormal cortical mRNA expression of key receptors involved in synaptic transmission and neuronal growth and development, changes that were consistent with the electrophysiological and morphologic changes. These findings demonstrate the lasting sequelae of perinatal fentanyl exposure on sensory processing and function.SIGNIFICANCE STATEMENT This is the first study to show that exposure to fentanyl in the womb results in behavioral, circuitry, and synaptic effects that last at least to adolescence. We also show, for the first time, that this exposure has different, lasting effects on synapses in different cortical areas.

Effects of the estrous cycle and ovarian hormones on central expression of interleukin-1 evoked by stress in female rats.

Exposure to stressors such as foot shock (FS) leads to increased expression of multiple inflammatory factors, including the proinflammatory cytokine interleukin-1 (IL-1) in the brain. Studies have indicated that there are sex differences in stress reactivity, suggesting that the fluctuations in gonadal steroid levels across the estrous cycle may play a regulatory role in the stress-induced cytokine expression. The present studies were designed to investigate the role of 17-ß-estradiol (E2) and progesterone (Pg) in regulating the cytokine response within the paraventricular nucleus (PVN) of the hypothalamus through analysis of gene expression with real-time RT-PCR. Regularly cycling female rats showed a stress-induced increase in PVN IL-1 levels during the diestrous, proestrous, and estrous stages. During the metestrous stage, no change in IL-1 levels was seen following FS; however, estrogen receptor (ER)-ß levels did increase. Ovariectomy resulted in an increase in PVN IL-1 levels, which was attenuated by treatment with estradiol benzoate (10 or 50 µg), indicating an E2-mediated anti-inflammatory effect. Ovariectomized rats treated with Pg (500 or 1,250 µg) showed no alteration in IL-1 levels, but Pg did up-regulate ER-ß gene expression. The results from the current study implicate a potential mechanism through which high availability of endogenous Pg during the metestrous stage increases ER-ß sensitivity, which in turn attenuates the PVN IL-1 response to stress. Thus, the interaction between gonadal steroid hormones and their central receptors may exert a powerful inhibitory effect on neuroimmune consequences of stress throughout the estrous cycle.

Adolescent environmental enrichment induces social resilience and alters neural gene expression in a selectively bred rodent model with anxious phenotype.

Stress is a major influence on mental health status; the ways that individuals respond to or copes with stressors determine whether they are negatively affected in the future. Stress responses are established by an interplay between genetics, environment, and life experiences. Psychosocial stress is particularly impactful during adolescence, a critical period for the development of mood disorders. In this study we compared two established, selectively-bred Sprague Dawley rat lines, the "internalizing" bred Low Responder (bLR) line versus the "externalizing" bred High Responder (bHR) line, to investigate how genetic temperament and adolescent environment impact future responses to social interactions and psychosocial stress, and how these determinants of stress response interact. Male bLR and bHR rats were exposed to social and environmental enrichment in adolescence prior to experiencing social defeat and were then assessed for social interaction and anxiety-like behavior. Adolescent enrichment caused rats to display more social interaction, as well as nominally less social avoidance, less submission during defeat, and resilience to the effects of social stress on corticosterone, in a manner that seemed more notable in bLRs. For bHRs, enrichment also caused greater aggression during a neutral social encounter and nominally during defeat, and decreased anxiety-like behavior. To explore the neurobiology underlying the development of social resilience in the anxious phenotype bLRs, RNA-seq was conducted on the hippocampus and nucleus accumbens, two brain regions that mediate stress regulation and social behavior. Gene sets previously associated with stress, social behavior, aggression and exploratory activity were enriched with differential expression in both regions, with a particularly large effect on gene sets that regulate social behaviors. Our findings provide further evidence that adolescent enrichment can serve as an inoculating experience against future stressors. The ability to induce social resilience in a usually anxious line of animals by manipulating their environment has translational implications, as it underscores the feasibility of intervention strategies targeted at genetically vulnerable adolescent populations.

Adolescent environmental enrichment induces social resilience and alters neural gene expression in a selectively bred rodent model with anxious phenotype.

Stress is a major influence on mental health status; the ways that individuals respond to or copes with stressors determine whether they are negatively affected in the future. Stress responses are established by an interplay between genetics, environment, and life experiences. Psychosocial stress is particularly impactful during adolescence, a critical period for the development of mood disorders. In this study we compared two established, selectively-bred Sprague Dawley rat lines, the "internalizing" bred Low Responder (bLR) line versus the "externalizing" bred High Responder (bHR) line, to investigate how genetic temperament and adolescent environment impact future responses to social interactions and psychosocial stress, and how these determinants of stress response interact. Male bLR and bHR rats were exposed to social and environmental enrichment in adolescence prior to experiencing social defeat and were then assessed for social interaction and anxiety-like behavior. Adolescent enrichment caused rats to display more social interaction, as well as nominally less social avoidance, less submission during defeat, and resilience to the effects of social stress on corticosterone, in a manner that seemed more notable in bLRs. For bHRs, enrichment also caused greater aggression during a neutral social encounter and nominally during defeat, and decreased anxiety-like behavior. To explore the neurobiology underlying the development of social resilience in the anxious phenotype bLRs, RNA-seq was conducted on the hippocampus and nucleus accumbens, two brain regions that mediate stress regulation and social behavior. Gene sets previously associated with stress, social behavior, aggression and exploratory activity were enriched with differential expression in both regions, with a particularly large effect on gene sets that regulate social behaviors. Our findings provide further evidence that adolescent enrichment can serve as an inoculating experience against future stressors. The ability to induce social resilience in a usually anxious line of animals by manipulating their environment has translational implications, as it underscores the feasibility of intervention strategies targeted at genetically vulnerable adolescent populations.

Sporadic ALS with compound heterozygous mutations in the SQSTM1 gene.

Accumulating evidence suggests that heterozygous mutations in the SQSTM1 gene, which encodes p62 protein, are associated with amyotrophic lateral sclerosis (ALS). Here, we report a Japanese patient with sporadic, late-onset ALS who harbored compound heterozygous SQSTM1 mutations (p.[Val90Met];[Val153Ile]). Autopsy examination revealed that although TDP-43 pathology was rather widespread, the selective occurrence of p62-positive/TDP-43-negative cytoplasmic inclusions in the lower motor neurons (LMNs) was a characteristic feature. No Bunina bodies were found. Ultrastructurally, p62-positive cytoplasmic inclusions observed in the spinal anterior horn cells were composed of aggregates of ribosome-like granules and intermingled bundles of filamentous structures. Another feature of interest was concomitant Lewy body pathology. The occurrence of distinct p62 pathology in the LMNs in this patient indicates the pathogenic role of SQSTM1 mutations in the development of a subset of ALS.

Prognosis in patients with non-small cell lung cancer who received erlotinib treatment and subsequent dose reduction due to skin rash.

BACKGROUND: Severe skin rash as toxicity of erlotinib has been reported in relation to better response and survival. However, some patients require dose reduction due to skin toxicities, and their prognosis remains uncertain. We retrospectively evaluated the clinical course of non-small cell lung cancer patients receiving erlotinib at a reduced dose because of skin rash. PATIENTS AND METHODS: Among 76 patients treated with erlotinib, 55 patients who developed skin rash severer than grade 2 were divided into 2 groups: 24 patients treated with erlotinib with dose reduction because of skin rash (dose reduction group) and 31 patients without any dose reduction (non-dose reduction group). RESULTS: The median progression-free survival in the dose reduction and non-dose reduction groups was 341 and 70 days, respectively, and the median overall survival was 566 and 202 days, respectively (p < 0.001). In the dose reduction group, no smoking history, female sex, epidermal growth factor receptor gene mutation, and grade 3 skin rash were significant baseline factors. CONCLUSIONS: Patients who received erlotinib at a reduced dose following skin rash showed better survival than those without reduction. In cases of intolerable skin rash, patients may benefit from continuous treatment with a reduced dose of erlotinib.

Patterns of cognitive decline and somatosensory processing in a mouse model of amyloid accumulation.

Pain and cognitive decline increase with age. In particular, there is a troubling relationship between dementia and pain, with some studies showing higher prevalence and inadequate treatment of pain in this population. Alzheimer's disease (AD) is one of the most common causes of dementia in older adults. Amyloid plaques are a hallmark of AD. The downstream processes these plaques promote are believed to affect neuronal and glial health and activity. There is a need to better understand how the neuropathological changes of AD shape neural activity and pain sensitivity. Here, we use the 5XFAD mouse model, in which dense amyloid accumulations occur at early ages, and in which previous studies reported signs of cognitive decline. We hypothesized that 5XFAD mice develop sensory and pain processing dysfunctions. Although amyloid burden was high throughout the brain, including in regions involved with sensory processing, we identified no functionally significant differences in reflexive or spontaneous signs of pain. Furthermore, expected signs of cognitive decline were modest; a finding consistent with variable results in the literature. These data suggest that models recapitulating other pathological features of Alzheimer's disease might be better suited to studying differences in pain perception in this disease.

Divergent profiles of fentanyl withdrawal and associated pain in mice and rats.

Opioid abuse has devastating effects on patients, their families, and society. Withdrawal symptoms are severely unpleasant, prolonged, and frequently hinder recovery or lead to relapse. The sharp increase in abuse and overdoses arising from the illicit use of potent and rapidly-acting synthetic opioids, such as fentanyl, highlights the urgency of understanding the withdrawal mechanisms related to these drugs. Progress is impeded by inconsistent reports on opioid withdrawal in different preclinical models. Here, using rats and mice of both sexes, we quantified withdrawal behaviors during spontaneous and naloxone-precipitated withdrawal, following two weeks of intermittent fentanyl exposure. We found that both mice and rats lost weight during exposure and showed increased signs of distress during spontaneous and naloxone precipitated withdrawal. However, these species differed in their expression of withdrawal associated pain, a key contributor to relapse in humans. Spontaneous or ongoing pain was preferentially expressed in rats in both withdrawal conditions, while no change was observed in mice. In contrast, withdrawal associated thermal hyperalgesia was found only in mice. These data suggest that rats and mice diverge in how they experience withdrawal and which aspects of the human condition they most accurately model. These differences highlight each species' strengths as model systems and can inform experimental design in studies of opioid withdrawal.

Sickness-related odor communication signals as determinants of social behavior in rat: a role for inflammatory processes.

Infected animals are avoided by conspecifics, suggesting that the inflammatory cascade may play a significant role in odor communication. Injection of male rats with the bacterial mimetic, lipopolysaccharide (LPS, 100 microg/kg, i.p.), decreased investigation through a wire-mesh partition between healthy male partners. This avoidance response was observed in adult males in response to soiled bedding collected from sick rats, regardless of whether LPS was injected peripherally (100 microg/kg, i.p.) or centrally (0.25 or 2.5 microg, icv). The release of sickness-related odor cues was dose-dependently blocked by icv infusion of the anti-inflammatory cytokine, interleukin-10 (IL-10; 20 or 200 ng), and reproduced by icv infusion of pro-inflammatory cytokine, IL-1beta (5 or 50 ng). Subcutaneous pretreatment with either estradiol benzoate (20 microg/kg) or testosterone propionate (50 or 500 microg/kg) to adult males that were administered LPS inhibited release of aversive odor cues, but these hormones alone did not influence odor properties. Importantly, the avoidance response to sickness-related odor was not associated with changes in plasma corticosterone, testosterone, or IL-6 levels of odor donors. However, plasma IL-1beta concentrations of sick animals was in fact predictive of aversive responses in conspecifics, suggesting that the inflammatory cascade, but not plasma steroid hormones, is likely to mediate aversive properties in odor that functions to signal illness state to conspecifics.

A Neural System that Represents the Association of Odors with Rewarded Outcomes and Promotes Behavioral Engagement.

Odors are well known to elicit strong emotional and behavioral responses that become strengthened throughout learning, yet the specific cellular systems involved in odor learning and the direct influence of these on behavior are unclear. Here, we investigate the representation of odor-reward associations within two areas recipient of dense olfactory input, the posterior piriform cortex (pPCX) and the olfactory tubercle (OT), using electrophysiological recordings from mice engaged in reward-based learning. Neurons in both regions represent conditioned odors and do so with similar information content, yet the proportion of neurons recruited by conditioned rewarded odors and the magnitudes and durations of their responses are greater in the OT. Using fiber photometry, we find that OT D1-type dopamine-receptor-expressing neurons flexibly represent odors based on reward associations, and using optogenetics, we show that these neurons influence behavioral engagement. These findings contribute to a model whereby OT D1 neurons support odor-guided motivated behaviors.

Scent marking behavior as an odorant communication in mice.

In rodents, where chemical signals play a particularly important role in determining intraspecies interactions including social dominance and intersexual relationships, various studies have shown that behavior is sensitive to conspecific odor cues. Mice use urinary scent marks for communication with individual conspecifics in many social contexts. Urinary scent involves genetic information about individuals such as species, sex, and individual identity as well as metabolic information such as social dominance, and reproductive and health status, which are mediated by chemical proteins in scent marks including the major histocompatibility complex and the major urinary proteins. The odor of the predator which can be considered to be a threatening signal for the prey also modulate mouse behavior in which scent marking is suppressed in response to the cat odor exposure in mice. These odorant chemicals are detected and recognized through two olfactory bulbs, the role of which in detection of chemosignals with biological relevant appears to be differential, but partly overlapped. Mice deposit scent marks toward conspecifics to maintain their social relationships, and inhibit scent marking in a context where natural predator, cat odor is contained. This suppression of scent marking is long-lasting (for at least 7 days) and context-dependent, while the odorant signaling to conspecifics tends to appear frequently (over 24h but less than 7 days intervals) depending on the familiarity of each signal-recipient. It has been discussed that scent marking is a communicative behavior associated with territoriality toward conspecifics, indicating that the social signaling within species are sensitive to predator odor cues in terms of vulnerability to predation risk.

A new test paradigm for social recognition evidenced by urinary scent marking behavior in C57BL/6J mice.

Olfaction is a major sensory element in intraspecies recognition and communication in mice. The present study investigated scent marking behaviors of males of the highly inbred C57BL/6J (C57) strain in order to evaluate the ability of these behaviors to provide clear and consistent measures of social familiarity and response to social signals. C57 males engage in scent marking when placed in a chamber with a wire mesh partition separating them from a conspecific. Male mice (C57 or outbred CD-1 mice) showed rapid habituation of scent marking (decreased marking over trials) with repeated exposure at 24-h intervals, to a stimulus animal of the C57 or CD-1 strains, or to an empty chamber. Subsequent exposure to a genetically different novel mouse (CD-1 after CD-1 exposure, or CD-1 after C57 exposure) or to a novel context (different shaped chamber) produced recovery of marking, while responses to a novel but genetically identical mouse (C57 after C57 exposure) or to the empty chamber did not. This finding demonstrated that male mice differentiate familiar and novel conspecifics as expressed by habituation and recovery of scent marking, but neither C57 or CD-1 mice can differentiate new vs. familiar C57 males; likely due to similarities in their odor patterns. The data also indicate that scent marking can differentiate novel from familiar contexts.

Inter- and intra-mouse variability in odor preferences revealed in an olfactory multiple-choice test.

Animals choose between sensory stimuli, a highly complex behavior which includes detection, discrimination, preference, and memory processes. Rodents are reported to display robust preferences for some odors, for instance, in the context of choosing among possible mates or food items. In contrast to the apparent robustness of responses toward these and other "ethologically relevant" odors, little is known about the robustness of behaviors toward odors which have no overt role in the rodent ecological niche, so-called "nonethologically relevant" odors. We developed an apparatus for monitoring the nose-poking behavior of mice and used this apparatus to explore the prevalence and stability of choices among different odors both across mice, and within mice over successive days. Mice were tested with a panel of either ethologically relevant or nonethologically relevant odors in an olfactory multiple-choice test. Significant preferences to nonethologically relevant odors were observed across the population of mice, with longer investigation durations to some odors more than to others. However, we found substantial inter-mouse variability in these responses, and that responses to these odors even varied within mice across days of testing. Tests with ethologically relevant odors revealed that responses toward these odors were also variable across mice, but within individual mice, responses were somewhat stable. This work establishes an olfactory multiple-choice test for monitoring odor investigation, choice, and preference behaviors and the application of this apparatus to assess across- and within-mouse odor-preference choice stability. These results highlight that odor preferences, as assayed by measuring choice behaviors, are variable. (PsycINFO Database Record

Scent marking behavior in male C57BL/6J mice: sexual and developmental determination.

The present study investigated urinary scent marking behavior in male C57BL/6J (C57) mice as olfactory social signaling. In Experiment 1, when compared scent marking toward adult males, C57 males showed substantial scent marking toward CD-1 males and even toward the odor alone of CD-1 males, but not toward C57 males. Experiment 2 explored scent marking in C57 males of different ages to males and females, and juveniles and adults of the same strain. C57 males deposited more marks than control conditions only toward an adult C57 female when tested at 100 days of age, but not at 60 days of age. Development of urine marking behavior was investigated in C57 males at the ages of 30, 60, 90, and 120 days in Experiment 3. When tested alone (control) or confronted with a C57 male, C57 males showed diminished scent marks throughout development. Compared to controls, marking toward a CD-1 male increased after the age of 60 days, while marks toward an adult female showed significant increases after the age of 90 days. This difference in scent marking depending on the sex of the stimulus animal is likely to be associated with development of sexual behavior, in which males need to set up territories against other males prior to advertising to females. Although highly inbred strains have similar odor components, C57 males are able to detect and deposit urine marks after puberty as social communication depending on age, sex, and genetic differences in the opponents.

The role of neuroinflammation in the release of aversive odor cues from footshock-stressed rats: Implications for the neural mechanism of alarm pheromone.

Stressed animals have been known to release aversive chemosignals toward which conspecifics show avoidance-like responses. The present studies assessed whether inflammatory cytokine responses provoked by footshock stress modulate odor signals released from male rats. Male rats were exposed to 30min of intermittent footshock (60 shocks, 1.0mA, 100ms each, variable ITI of 30s) or remained in their home cages as non-stressed controls. Real time RT-PCR analysis of brain tissues indicated that footshock increased the pro-inflammatory cytokine, IL-1ß and hnCRH as well as c-fos mRNA expressions in the paraventricular nucleus, and the bed nucleus of the stria terminalis, and increased plasma corticosterone levels. Soiled bedding collected from rats exposed to 30-min, but not 5-min, of footshock elicited a differential response, as expressed by decreased sniffing and increased avoidance in male test subjects. Soiled bedding from rats given corticosterone injection (s.c. 1.25 or 3.75mg/ml) 3h before bedding collection evoked no avoidance response in odor-recipients. Furthermore, ICV infusion of the anti-inflammatory cytokine IL-10 (20 or 200ng) into the stimulus animals 30-min before a 30-min footshock session, had no effect on plasma corticosterone levels in the stimulus animals, but attenuated the release of aversive odor as indicated by dose-dependently diminished avoidance in odor-recipient rats. These results demonstrated that stressed rats release odorant cues that cause other rats to move away from the source of the signal. Such stress-induced chemosignals may be mediated by inflammatory cytokine responses in the brain.

A patient with fragile x-associated tremor/ataxia syndrome presenting with executive cognitive deficits and cerebral white matter lesions.

Fragile X-associated tremor/ataxia syndrome (FXTAS) is a late-onset neurodegenerative disorder that primarily affects males who are carriers of a premutation of a CGG expansion in the FMR1 gene. In Asian populations, FXTAS has rarely been reported. Here, we report the case of a Japanese FXTAS patient who showed predominant executive cognitive deficits as the main feature of his disease. In contrast, the patient exhibited only very mild symptoms of intention tremor and ataxia, which did not interfere with daily activities. A gene analysis revealed that the patient carried a premutation of a CGG expansion (111 CGG repeats) in the FMR1 gene. The mRNA expression level of FMR1 in the patient was 1.5-fold higher than in controls. On brain MRI scans, fluid-attenuated inversion recovery images showed high-intensity lesions in the middle cerebellar peduncles and the cerebral white matter, with a frontal predominance. The present case extends previous notions regarding the cognitive impairment in FXTAS patients. Recognizing FXTAS patients with predominant cognitive impairment from various ethnic backgrounds would contribute to our understanding of the phenotypic variation of this disease.

Acute illness induces the release of aversive odor cues from adult, but not prepubertal, male rats and suppresses social investigation by conspecifics.

The present study examined odorant communication during acute illness provoked by injection of lipopolysaccharide (LPS; 100 microg/kg) and how these effects vary between prepubertal and adult conspecifics. Exposure to odor of LPS-treated adult male rats produced increased avoidance in both sexes of adults and prepubertal male partners. This response was not found when they were exposed to odor of LPS-treated prepubertal males. Even a 2.5-fold higher load of LPS in prepubertal males failed to produce aversive odor cues, suggesting that the difference in the odor is not a simple issue of dose/body volume. Both estradiol benzoate (20 microg/kg) and testosterone propionate (500 microg/kg), but not dihydrotestosterone (500 microg/kg) pretreatment in prepubertal males administered LPS restored the expression of aversive odor. These hormone treatments per se did not influence odor properties of prepubertal males, indicating that estrogen receptors may play a key regulatory role in the expression of aversive odor in LPS-treated prepubertal rats. These data suggest that the expression of sickness-related odor emerges through puberty, and likely involves a complex interaction between inflammation and sex steroids across development.

Social features of scent-donor mice modulate scent marking of C57BL/6J recipient males.

Territorial male mice can form familiarity-dependent amicable relationships, suggesting that they manage their territorial aggressiveness based on individual recognition, which may be mediated by olfactory signals. The present study demonstrated modulatory effect of odorant cues from mice of different social/housing groups on territorial scent marking by C57BL/6J males. Pair-housed males deposited few scent marks in a novel situation without mouse odors, while singly-housed males marked more in the same situation (Experiment 1). However, when confronted by a conspecific, singly-housed males made fewer marks to pair-housed than to singly-housed stimulus males, while pair-housed males showed few marks to either stimulus animal (Experiment 2). Reduction in scent marking in singly-housed males was also seen on exposure to urine scent alone from a pair-housed male, indicating that the inhibitory cue is mediated by urinary odor (Experiment 3). This inhibitory odor was effective even when singly-housed males were placed in a no-odor environment following exposure to soiled bedding used by pair-housed males (Experiment 4). When singly-housed males were exposed to scent from subordinate males, they showed less marking than when the stimulus scent was from dominants or singly-housed males (Experiment 5). Scent marking was not influenced by the total amount of urine scent marks deposited on the substrates (Experiment 6), suggesting that it is a particular component in urinary odor rather than the magnitude of the odor, that plays a significant role in recipient behavior. Together, these experiments indicate that conspecific male odor signals modulate territorial scent marking behavior in mice.

Correlation between pharmacological efficacy of cyclosporine A and tacrolimus, evaluated by lymphocyte immunosuppressant-sensitivity test (LIST) with MTT assay procedure in renal transplant recipients.

The dose of calcineurin inhibitors in renal transplantation has been adjusted, based on the therapeutic drug monitoring data. However, the data do not always correlate with clinical drug efficacy. In vitro response of peripheral-blood mononuclear cells to immunosuppressive drugs is reported to correlate with the recipient-response to therapeutic efficacy of the drug. We report, here, usefulness of a lymphocyte immunosuppressant sensitivity test for the estimation of individual drug sensitivity in renal transplant recipients. The LIST we have developed includes MTT assay procedures without the use of radioisotope-labeled compounds, which is convenient for general hospital use. Utilizing this procedure, we compared the pharmacological efficacy between cyclosporine A and tacrolimus in 41 renal transplant recipients.