Hitomi Sakano

Odorant stimulation enhances survival of olfactory sensory neurons via MAPK and CREB

Olfactory sensory neurons (OSNs) can be sensitized to odorants by repeated exposure, suggesting that an animals responsiveness to olfactory cues can be enhanced at the initial stage of detection. However, because OSNs undergo a regular cycle of apoptosis and replacement by ostensibly naive, precursor-derived neurons, the advantage of sensitization would be lost in the absence of a mechanism for odorant-enhanced survival of OSNs. Using recombinant adenoviruses in conjunction with surgical and electrophysiological techniques, we monitored OSN survival and function in vivo and find that odorant exposure selectively rescues populations of OSNs from apoptosis. We further demonstrate that odorant stimuli rescue OSNs in a cAMP-dependent manner by activating the MAPK/CREB-dependent transcriptional pathway, possibly as a result of expression of Bcl-2.

Characterization of the influence of polyol fatty acid esters on the permeation of diclofenac through rat skin.

In this study, we investigated the effect of sefsols on the skin to clarify the mechanism of the sefsol enhancement effect. In vitro percutaneous absorption experiments were performed using Franz diffusion cells. Removal of the stratum corneum and delipidization of the skin increased the permeation of diclofenac from aqueous suspension, with the enhancement effects being similar for both treatments. Further enhancement effects of diclofenac permeation by sefsol through the stripped and delipidized skin were not observed. Attenuated total reflectance-Fourier transform infrared spectroscopy (ATR-FTIR) was employed to investigate the biophysical changes in the stratum corneum lipids by sefsols. One of the sefsols, propylene glycol mono caprylate (S-218), induced higher and broader absorbance shifts in both asymmetric and symmetric C-H bond stretching regions. However, no significant differences were observed among the sefsols with respect to peak heights and areas for both absorbances when compared with H2O treatment. These results suggest that sefsol may change the lipid-chain fluidity of the stratum corneum without lipid extraction. The accumulated amounts of diclofenac in the skin significantly increased in the presence of sefsol. Also, the amounts of diclofenac in the skin increased with the amount of sefsol in the skin. This sefsol enhancement effect was reversed at 12 h after treatment. Thus, enhancement of the diclofenac flux by sefsols is reversible and may be due to a change in the lipid-chain fluidity of the stratum corneum and improvement in drug partitioning to the stratum corneum.

Effect of Fatty Acid Diesters on Permeation of Anti-Inflammatory Drugs Through Rat Skin

ABSTRACT Four fatty acid diesters (diethyl succinate, diethyl adipate, diethyl sebacate, and diisopropyl adipate) were used to study their enhancement effect on the permeation of four non-steroidal anti-inflammatory drugs (NSAIDs: ketoprofen, indomethacin, diclofenac sodium, and ibuprofen) through rat abdominal skin. With the diester pretreatment, drug permeation increased and the lag times decreased. No relationship was observed between the solubilities of the drugs in the diesters and the diester enhancement effects. The enhancement effect decreased with an increase of the drug lipophilicity, but increased with an increase of the lipophilic index of the diester up to about 3.5, after which the enhancement effect decreased or remained constant. Attenuated total reflectance-Fourier transform infrared spectroscopy (ATR-FTIR) was employed to investigate the biophysical changes in the stratum corneum lipids caused by the diesters. The FTIR results showed that treatment of the skin with diesters did not produce a blue shift in the asymmetric and symmetric C–H stretching peak positions. However, all of the above diesters showed a decrease in peak heights and areas for both asymmetric and symmetric C–H stretching absorbances in comparison with water treatment. These results suggested that the diesters were more effective for enhancing the penetration of hydrophilic drugs than lipophilic drugs, and the enhancing effect of lipophilic diesters was more effective than that of hydrophilic diesters. The enhancement effects of diesters may be due to their causing lipid extraction in the skin.

Intense and specialized dendritic localization of the fragile X mental retardation protein in binaural brainstem neurons: a comparative study in the alligator, chicken, gerbil, and human.

Neuronal dendrites are structurally and functionally dynamic in response to changes in afferent activity. The fragile X mental retardation protein (FMRP) is an mRNA binding protein that regulates activity‐dependent protein synthesis and morphological dynamics of dendrites. Loss and abnormal expression of FMRP occur in fragile X syndrome (FXS) and some forms of autism spectrum disorders. To provide further understanding of how FMRP signaling regulates dendritic dynamics, we examined dendritic expression and localization of FMRP in the reptilian and avian nucleus laminaris (NL) and its mammalian analogue, the medial superior olive (MSO), in rodents and humans. NL/MSO neurons are specialized for temporal processing of low‐frequency sounds for binaural hearing, which is impaired in FXS. Protein BLAST analyses first demonstrate that the FMRP amino acid sequences in the alligator and chicken are highly similar to human FMRP with identical mRNA‐binding and phosphorylation sites, suggesting that FMRP functions similarly across vertebrates. Immunocytochemistry further reveals that NL/MSO neurons have very high levels of dendritic FMRP in low‐frequency hearing vertebrates including alligator, chicken, gerbil, and human. Remarkably, dendritic FMRP in NL/MSO neurons often accumulates at branch points and enlarged distal tips, loci known to be critical for branch‐specific dendritic arbor dynamics. These observations support an important role for FMRP in regulating dendritic properties of binaural neurons that are essential for low‐frequency sound localization and auditory scene segregation, and support the relevance of studying this regulation in nonhuman vertebrates that use low frequencies in order to further understand human auditory processing disorders. J. Comp. Neurol. 522:2107–2128, 2014.

Influence of pH on the permeability of p-toluidine and aminopyrine through shed snake skin as a model membrane.

The influence of pH on the permeability of p-toluidine (pKa, 5.3) and aminopyrine (pKa, 5.0) through shed snake skin as a model membrane was studied. The pH was adjusted to several values, and the solubility of the drugs in each donor was measured. Flux rates and permeability coefficients were calculated from the steady-state penetration portions. The flux rates of p-toluidine decreased as the pH value in the donor solution increased. On the other hand, the flux rates of aminopyrine were constant at any pH value. The permeability coefficients of each drug increased as the pH value in the donor solution increased. The partition coefficients (octanol/buffer) of each drug were dependent on the molecular fraction of un-ionized species. From these results, it is suggested that ionized species of p-toluidine transports through shed snake skin, but the ionized species of aminopyrine does not.

Proteomic analyses of nucleus laminaris identified candidate targets of the fragile X mental retardation protein

The avian nucleus laminaris (NL) is a brainstem nucleus necessary for binaural processing, analogous in structure and function to the mammalian medial superior olive. In chickens (Gallus gallus), NL is a well‐studied model system for activity‐dependent neural plasticity. Its neurons have bipolar extension of dendrites, which receive segregated inputs from two ears and display rapid and compartment‐specific reorganization in response to unilateral changes in auditory input. More recently, fragile X mental retardation protein (FMRP), an RNA‐binding protein that regulates local protein translation, has been shown to be enriched in NL dendrites, suggesting its potential role in the structural dynamics of these dendrites. To explore the molecular role of FMRP in this nucleus, we performed proteomic analysis of NL, using micro laser capture and liquid chromatography tandem mass spectrometry. We identified 657 proteins, greatly represented in pathways involved in mitochondria, translation and metabolism, consistent with high levels of activity of NL neurons. Of these, 94 are potential FMRP targets, by comparative analysis with previously proposed FMRP targets in mammals. These proteins are enriched in pathways involved in cellular growth, cellular trafficking and transmembrane transport. Immunocytochemistry verified the dendritic localization of several proteins in NL. Furthermore, we confirmed the direct interaction of FMRP with one candidate, RhoC, by in vitro RNA binding assays. In summary, we provide a database of highly expressed proteins in NL and in particular a list of potential FMRP targets, with the goal of facilitating molecular characterization of FMRP signaling in future studies.

Intense and specialized dendritic localization of the fragile X mental retardation protein in binaural brainstem neurons: A comparative study in the alligator, chicken, gerbil, and human: FMRP localization in NL/MSO dendrites

Neuronal dendrites are structurally and functionally dynamic in response to changes in afferent activity. The fragile X mental retardation protein (FMRP) is an mRNA binding protein that regulates activity‐dependent protein synthesis and morphological dynamics of dendrites. Loss and abnormal expression of FMRP occur in fragile X syndrome (FXS) and some forms of autism spectrum disorders. To provide further understanding of how FMRP signaling regulates dendritic dynamics, we examined dendritic expression and localization of FMRP in the reptilian and avian nucleus laminaris (NL) and its mammalian analogue, the medial superior olive (MSO), in rodents and humans. NL/MSO neurons are specialized for temporal processing of low‐frequency sounds for binaural hearing, which is impaired in FXS. Protein BLAST analyses first demonstrate that the FMRP amino acid sequences in the alligator and chicken are highly similar to human FMRP with identical mRNA‐binding and phosphorylation sites, suggesting that FMRP functions similarly across vertebrates. Immunocytochemistry further reveals that NL/MSO neurons have very high levels of dendritic FMRP in low‐frequency hearing vertebrates including alligator, chicken, gerbil, and human. Remarkably, dendritic FMRP in NL/MSO neurons often accumulates at branch points and enlarged distal tips, loci known to be critical for branch‐specific dendritic arbor dynamics. These observations support an important role for FMRP in regulating dendritic properties of binaural neurons that are essential for low‐frequency sound localization and auditory scene segregation, and support the relevance of studying this regulation in nonhuman vertebrates that use low frequencies in order to further understand human auditory processing disorders. J. Comp. Neurol. 522:2107–2128, 2014.

Intense and specialized dendritic localization of the fragile X mental retardation protein in binaural brainstem neurons

Neuronal dendrites are structurally and functionally dynamic in response to changes in afferent activity. The fragile X mental retardation protein (FMRP) is an mRNA binding protein that regulates activity‐dependent protein synthesis and morphological dynamics of dendrites. Loss and abnormal expression of FMRP occur in fragile X syndrome (FXS) and some forms of autism spectrum disorders. To provide further understanding of how FMRP signaling regulates dendritic dynamics, we examined dendritic expression and localization of FMRP in the reptilian and avian nucleus laminaris (NL) and its mammalian analogue, the medial superior olive (MSO), in rodents and humans. NL/MSO neurons are specialized for temporal processing of low‐frequency sounds for binaural hearing, which is impaired in FXS. Protein BLAST analyses first demonstrate that the FMRP amino acid sequences in the alligator and chicken are highly similar to human FMRP with identical mRNA‐binding and phosphorylation sites, suggesting that FMRP functions similarly across vertebrates. Immunocytochemistry further reveals that NL/MSO neurons have very high levels of dendritic FMRP in low‐frequency hearing vertebrates including alligator, chicken, gerbil, and human. Remarkably, dendritic FMRP in NL/MSO neurons often accumulates at branch points and enlarged distal tips, loci known to be critical for branch‐specific dendritic arbor dynamics. These observations support an important role for FMRP in regulating dendritic properties of binaural neurons that are essential for low‐frequency sound localization and auditory scene segregation, and support the relevance of studying this regulation in nonhuman vertebrates that use low frequencies in order to further understand human auditory processing disorders. J. Comp. Neurol. 522:2107–2128, 2014.