A novel localization of molt-inhibiting hormone in the tegumental glands of shrimp Penaeus monodon and its possible role in shrimp molting

• Molt-inhibiting hormone (MIH) was found to be stored in the tegumental glands located in the integument and at the retina of the eyestalk, these are the first report.
• Localization of MIH at the tegumental gland located in the integument and its distribution all over the epidermal cells evidenced the new route of molt regulating mechanism in which it probably regulates the molting at the target organs by disrupting the conversion of ecdysteroids.
• The synthesis and release of MIH from X-organ/sinus gland during the molt cycle were the continual processes and showing intermittent secretion; there was no obviously change at any molt stage.
• Localization of MIH at the tegumental gland located in the retina suggests its function possibly relate to the vision of shrimp and could link to the molting behavior.

Molt-inhibiting hormone (MIH) is commonly localized in the X-organ sinus gland complex (XOSG) of the crustacean. The present study aimed to elucidate the expression of MIH at the subcellular and cellular levels in the eyestalks and integument of juvenile Penaeus monodon during the molting cycle. Gene expression of Pem-MIH1 in the optic lobes showed a single PCR product at 172 bp, and was restricted only to the eyestalk XOSG but not detected in pleopods, cephalothorax integument, muscle, hepatopancreas or retina. Quantitative analysis of gene expression in the eyestalks demonstrated no significant alteration of Pem-MIH1 mRNA in XOSG during the molt cycle. Immunohistochemistry using antibody against recombinant molt-inhibiting hormone-like peptide (anti-MIH-like) revealed variable staining intensities of individual MTXOs but were most persistently intense in SGs. During the molt cycle, the mean numbers of XO immunoreactive cells were slightly oscillated but not statistically different. The quantitative immunohistochemistry measured from XOSG illustrated minimal fluctuations of the values obtained suggesting the periodical synthesis and release of MIH from XOSG system during the molt cycle. The positive reaction was also detected in the tegumental glands located at the retina and the integument of walking leg, base of eyestalk and cephalothorax of the shrimp. The large variations of immunostaining and amount of reactive tegumental glands in the eyestalk were noted throughout the molt cycle. The numbers of the MIH-like immunoreactive glands obtained from the retina as well as the integument base did not significantly change during the molt cycle but tend to increase during postmolt to intermolt and decrease during premolt. These results suggest that the MIH-like peptides in the tegumental glands are probably released mostly during the late premolt. Our findings thus propose the novel storage site of MIH besides SG of the eyestalk including a new target tissue, an epidermal cell, the role of which may link to the complicated molting regulation and/or the dark-adaptation of shrimp.