Hideaki Inagaki

Enhancement of the acoustic startle reflex by an alarm pheromone in male rats

Recently, we reported that an alarm pheromone released from the perianal region of male rats aggravated stress-induced hyperthermia and increased defensive and risk assessment behaviors in recipient male rats. Based on these results, we hypothesized that the primary effect of the alarm pheromone is to increase anxiety; however, there is still no clear evidence for this pheromone effect. Therefore, we examined this issue by assessing the effect of the alarm pheromone on the acoustic startle reflex (ASR), which is a useful index for studying negative emotions such as anxiety in rats. The alarm pheromone enhanced the ASR for 105-dB auditory stimuli, but not for those of 90 and 120 dB, when these three intensities of sound were used randomly. The same results were obtained when one of these three intensities was used repeatedly. In addition, pretreatment with diazepam (i.p.) at doses of 0.7 and 2.0 mg/kg suppressed the ASR of the pheromone recipients, whereas the lower dose (0.2 mg/kg) slightly attenuated the pheromone effect and the control injection (vehicle) had no effect. These results indicate that the alarm pheromone enhances the ASR by increasing anxiety in recipient rats, suggesting that the primary effect of the alarm pheromone is to increase the anxiety level.

Opioid mediated suppressive effect of milk-derived lactoferrin on distress induced by maternal separation in rat pups

The present study assessed the effects of bovine milk-derived lactoferrin (bLf) on distress activities induced by maternal separation in 5- to 18-day-old rat pups. The rat pups were injected with BSA (100 mg/kg, i.p.; control) or bLf (100 mg/kg, i.p.) 30 min before the behavioral test. Distress activity was estimated by means of recording body movements or ultrasonic vocalizations (USVs). After 5 min of maternal separation, bLf significantly (P<0.01) suppressed body movements, particularly in the 10-day-old pups. This suppressive effect of bLf was reversed by pretreatment with naloxone, CTOP, and norBNI at doses of 0.1-1 mg/kg. Additionally, USVs were also suppressed by bLf, which was reversed by pretreatment with naloxone. Inhibition of nitric oxide synthase (NOS) with nitro-L-arginine methyl ester (L-NAME) dose dependently (3-10 mg/kg) suppressed separation-induced USV production in 10-day-old pups. Interestingly, the suppressive effect of bLf was completely reversed by pretreatment with a low dose (1 mg/kg) of L-NAME, which did not affect the USVs with single application. These findings demonstrate that milk-derived bLf suppresses distress induced by maternal separation via an opioid-mediated mechanism. Furthermore, bLf possibly activates NOS, and an elevated nitric oxide may cause some modification of the opioid system.

The volatility of an alarm pheromone in male rats

The volatility of an alarm pheromone in male rats. PHYSIOL BEHAV 00(0) 000-000, 2008. We previously reported that an alarm pheromone released from the perianal region of male rats is perceived by the vomeronasal organ and evokes stress-induced hyperthermia and defensive and risk assessment behavior. In addition, we recently reported that the alarm pheromone enhances the acoustic startle reflex (ASR). However, in contrast to our knowledge about such biological aspects of the pheromone, information concerning the physical character of the alarm pheromone is extremely limited. In this study, we investigated the volatility of the alarm pheromone using enhancement of the ASR as an index of the pheromone effect. The alarm pheromone enhanced the ASR when it was presented at a distance of 10 mm but not at 200 mm. In addition, the pheromone effect was observed even after the pheromone was trapped in the adsorbent (Tenax) and then extracted using purified water. These results suggest that the alarm pheromone is both volatile and water soluble.

The influence of social environmental condition on the production of stress-induced 22 kHz calls in adult male Wistar rats

Adult rats emit 22 kHz ultrasonic vocalizations (USVs) in response to aversive stimuli, and these sounds are suggested to have communicative information among conspecifics. It is conceivable that social environment during development of rats has relevance to the emission of 22 kHz USVs. To examine the effects of social environment after weaning on production of stress-induced USVs, we compared the amount of emission of USVs among three groups of rats reared under different conditions after weaning. One group of rats was housed individually, and the other two groups were housed in pairs, in which social hierarchy of the pair was determined by social dominance-subordination relationships. The USVs were induced by acute mild somatic stimuli on the back and neck. Individually reared rats emitted much fewer USVs than pair-reared rats. In addition, socially subordinate rats emitted more USVs compared with socially dominant ones. These results suggest that not only social interaction but also the status in social hierarchy may play an important role in the process of the development of USVs induced by somatic stimuli.

Identification of a pheromone that increases anxiety in rats

Significance Animals communicate with other members of the same species by using pheromones. Rats release a specific odor into the air when they are stressed. Because this stress-related odor increases anxiety in the other rats, it is likely that there are some anxiogenic molecules in this odor. In this study, we have tried to identify these molecules. We discovered that a combination of two molecules, 4-methylpentanal and hexanal, increased anxiety in rats. We concluded that the mixture is a pheromone in rats that can increase anxiety in conspecifics. Furthermore, we proposed that the neural circuit relating to anxiety was only activated when the two molecules were simultaneously perceived by two separate olfactory systems. Chemical communication plays an important role in the social lives of various mammalian species. Some of these chemicals are called pheromones. Rats release a specific odor into the air when stressed. This stress-related odor increases the anxiety levels of other rats; therefore, it is possible that the anxiety-causing molecules are present in the stress-related odorants. Here, we have tried to identify the responsible molecules by using the acoustic startle reflex as a bioassay system to detect anxiogenic activity. After successive fractionation of the stress-related odor, we detected 4-methylpentanal and hexanal in the final fraction that still possessed anxiogenic properties. Using synthetic molecules, we found that minute amounts of the binary mixture, but not either molecule separately, increased anxiety in rats. Furthermore, we determined that the mixture increased a specific type of anxiety and evoked anxiety-related behavioral responses in an experimental model that was different from the acoustic startle reflex. Analyses of neural mechanisms proposed that the neural circuit related to anxiety was only activated when the two molecules were simultaneously perceived by two olfactory systems. We concluded that the mixture is a pheromone that increases anxiety in rats. To our knowledge, this is the first study identifying a rat pheromone. Our results could aid further research on rat pheromones, which would enhance our understanding of chemical communication in mammals.

Didemnilactones A and B and neodidemnilactone, three new fatty acid metabolites isolated from the tunicate didemnum moseleyi (Herdman)

Abstract Three new fatty acid metabolites didemnilactones A (1) and B (2) and neodidemnilactone (3) were isolated from the tunicate Didemnum moseleyi (Herdman). Their structures including absolute stereochemistry were established on the basis of spectral studies and chemical synthesis. Didemnilactones exhibited inhibitory activity against lipoxygenase and weak binding activity to leukotriene B4 receptors.

Close relationship between the frequency of 22-kHz calls and vocal tract length in male rats

It is known that the size of the components of the sound production apparatus in mammals may affect the acoustic structure of vocalizations. Therefore, some acoustic variables such as voice frequency may change with age in association with body size and body weight increases. However, whether this relationship also applies to ultrasonic vocalizations emitted by laboratory rats has not been investigated. Thus, in the present study, we first recorded changes in three acoustic variables (mean frequency, duration, and bandwidth) of air puff-induced 22-kHz calls in male rats during their growth period and assessed the relationship between these changes and body weight gain (Experiment 1). Then we directly recorded several body size measures including components of the sound production apparatus in 6- and 12-week-old male rats and examined the correlation between these values and the acoustic variables of 22-kHz calls (Experiment 2). In Experiment 1, the mean frequency of 22-kHz calls in male rats during the growth period showed negative correlations with body weight gain, while the duration of 22-kHz calls showed positive correlations. In Experiment 2, only a close negative correlation between the mean frequency of 22-kHz calls and vocal tract length in male rats was found. These results suggest that the age-related decrease in the mean frequency of 22-kHz calls may be ascribed to anatomical elongation of the vocal tract length in association with the growth of male rats. These acoustic differences could inform the receivers about the age of the signaler.

The alarm pheromone in male rats as a unique anxiety model: psychopharmacological evidence using anxiolytics.

Previously, we demonstrated that an alarm pheromone released from male donor Wistar rats evoked anxiety-related physiological and behavioral responses in recipient rats. Thus, we believe that this pheromone may increase anxiety levels in rats. In the current study, we evaluated the predictive validity of this alarm pheromone-induced anxiogenic effect in detail by investigating whether six types of human anxiolytics, each of which has a different mechanism of action, were efficacious in reducing anxiety, using changes in the acoustic startle reflex (ASR) as an index. The alarm pheromone-enhanced ASR was not affected by vehicle pretreatment but was dose-dependently attenuated by pretreatment with midazolam, phenelzine, propranolol, clonidine, and CP-154,526-although not buspirone. These results may reflect some aspects of the predictive validity of the alarm pheromone-induced anxiety in rats as an animal model of human anxiety.

Didemnilactone and neodidemnilactone, two new fatty acid metabolites possessing a 10-membered lactone from the tunicate didemnum moseleyi (herdman)

Didemnilactone (1) and neodidemnilactone (2), two new fatty acid metabolites possessing a 10-membered lactone were isolated from the colonial tunicate Didemnum moseleyi (Herdman). Their structures including absolute stereochemistry were determined on the basis of spectral studies and the enantioselective synthesis of the antipode of 2.

Discovery, Optimization, and Pharmacological Characterization of Novel Heteroaroylphenylureas Antagonists of C-C Chemokine Ligand 2 Function

Through the application of TRAP (target-related affinity profiling), we identified a novel class of heteroaroylphenylureas that inhibit human CCL2-induced chemotaxis of monocytes/macrophages both in vitro and in vivo. This inhibition was concentration-dependent and selective with regard to other chemokines. The compounds, however, did not antagonize the binding of (125)I-labeled CCL2 to the CCR2 receptor nor did they block CCR2-mediated signal transduction responses such as calcium mobilization. Optimization of early leads for potency and pharmacokinetic parameters resulted in the identification of 17, a potent inhibitor of chemotaxis (IC(50) = 80 nM) with excellent oral bioavailability in rats (F = 60%). Compound 17 reduced swelling and joint destruction in two rat models of rheumatoid arthritis and delayed disease onset and produced near complete resolution of symptoms in a mouse model of multiple sclerosis.