Little is known about the regulation of the neonatal gastro-intestinal smooth muscle (SM) contraction. Screening of the proteom of the contractile machinery revealed that in the neonatal ileal SM, only the M133 isoform of the targeting subunit of the myosin phosphatase, MYPT1, is expressed whereas the adult ileum contains both the M133 and the M130 isoforms. Since expression of the different MYPT1 isoforms is known to be associated with different relaxing effects of PKA/PKG signaling we investigated whether the relaxing effect of cAMP/PKA and cGMP/PKG differs between longitudinal ileal SM from neonatal (< 1 day old) and adult (> 8 weeks) mice. In both adult and neonatal SM, 10 mM carbachol induced a tonic contraction, which in neonatal SM lacked the initial phasic overshoot. Addition of the stress related hormone urocortin (10 nM to 1 mM), known to signal through cAMP, at the plateau of the CCh-induced contraction induced a dose-dependent relaxation in the adult but not in the neonatal ileum. The dose response relation to forskolin and the PDE inhibitor theophylline was shifted to the right by more than one order of magnitude in the neonatal compared to the adult ileum (pIC50 values forskolin: 6.99 and 5.12 in adult and neonatal ileum respectively; theophyllin: 5.26 and 3.8 in adult and neonatal ileum respectively). Membrane permeable analogues of cAMP (db-cAMP, SP-cBIMPS) induced a dose-dependent relaxation only in adult SM strips. In contrast, membrane permeable cGMP analogues and the PDE5 inhibitor sildenalfil relaxed neonatal and adult ileal SM with similar potencies (sildenafil pIC50 adult SM 5.23 and neonatal SM 5.69). PKA/PKG signaling is known to decrease the Ca²⁺-sensitivity of contraction. Therefore we tested the Ca²⁺-desensitizing effect of cAMP and cGMP in ß-escin permeabilized ileal SM. In adult and neonatal SM, db-cAMP (0.3 mM) relaxed submaximally Ca2+-activated ileum by ~20%. In contrast 8-pCPT-cGMP (0.3 mM) relaxed neonatal SM by ~50% and adult SM by ~25%. In conclusion, neonatal ileal SM is more sensitive to cGMP/PKG than to cAMP/PKA signaling. This may in part be due to the higher degree of Ca²⁺-desensitization induced by cGMP compared to cAMP. Further, the higher degree of cGMP-induced Ca²⁺-desensitization in neonatal SM may be related to the different MYPT1 isoform expression.

Supported by Köln Fortune to F.E.