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ABSTRACT-The present study was conducted to establish a new mouse model of dry skin pruritus. The rostral back was treated daily with cutaneous application of acetone /ether (1:1) mixture (AE), water following AE (AEW), 1% sodium lauryl sulfate (SLS) or tape stripping (TS). On the day after 5-day treatment, although all four treatments significantly decreased stratum corneum (SC) hydration and increased transepidermal water loss (TEWL), only AEW treatment significantly increased spontaneous scratching. An increase in the frequency of TS produced the marked increase of TEWL, without significant effects on SC hydration and spontaneous scratching. In AEW-treated mice, changes in SC hydration and TEWL were marked in the initial 2-day period, while spontaneous scratching increased gradually from 3 days after starting the treatment. The degranulation of cutaneous mast cells was increased by SLS treatment but not by other treatments. There was no apparent difference in AEW-induced spontaneous scratching between mast cell-deficient mice (WBB6F1-W /W V ) and normal littermates (WBB6F1-+/+). Opioid antagonists, naloxone and naltrexone, (1 mg /kg, subcutaneously) significantly suppressed spontaneous scratching in AEW-treated mice. It is suggested that spontaneous scratching of AEW-treated mice is an itch-related response and a useful model for studying the mechanisms of dry skin pruritus.

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C3G is a guanine nucleotide exchange factor for Rap1 and is activated by the expression of Crk adaptor proteins. We found that expression of CrkI in COS cells induced significant tyrosine phosphorylation of C3G. To understand the mechanism by which C3G is phosphorylated and activated by Crk, we constructed a series of deletion mutants. Deletion of the amino terminus of C3G to amino acid 61 did not remarkably affect either tyrosine phosphorylation or Crk-dependent activation of C3G. When C3G was truncated to amino acid 390, C3G was still phosphorylated on tyrosine but was not effectively activated by CrkI. Deletion of the amino terminus of C3G to amino acid 579 significantly reduced the Crkdependent tyrosine phosphorylation of C3G and increased GTP-bound Rap1 irrespective of the presence of CrkI. We substituted all seven tyrosine residues in this region, amino acids 391-579, for phenylalanine for identification of the phosphorylation site. Among the substitution mutants, the C3G-Y504F mutant, in which tyrosine 504 was substituted by phenylalanine, was remarkably less activated and phosphorylated than the wild type. All the other substitution mutants were activated and tyrosyl-phosphorylated by the expression of CrkI. Thus, CrkI activates C3G by the phosphorylation of tyrosine 504, which represses the cis-acting negative regulatory domain outside the catalytic region.

Capsaicin has been suggested to act not only on thin primary afferents but also on the hypothalamus, but the neurotransmitter(s) of central capsaicin-sensitive neurons are unknown. The present study was conducted to determine whether any central, especially hypothalamic, glutamatergic terminals were sensitive to capsaicin. Capsaicin evoked glutamate release from slices of hypothalamus and lumbar dorsal horn, but not cerebellum. Such capsaicin action was Ca2+ dependent and inhibited by the capsaicin antagonist capsazepine. Vanilloid receptor subtype 1 mRNA was widely distributed in the brain, with a marked level in the hypothalamus and cerebellum, but not in the spinal cord. The results suggest that there are glutamatergic terminals sensitive to capsaicin in the hypothalamus.