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Effect of Phosphodiesterase Subtype3on the Cnp-Decreased Gastric Antral Smooth Muscle Spontaneous Contraction in Rats

Author: SunZuo
Tutor: CaiYingLan
School: Yanbian University
Course: Human Anatomy and Embryology
Keywords: CNP cAMP cGMP PDE antral circular smooth muscle spontaneous contraction
CLC: R333
Type: Master's thesis
Year: 2013
Downloads: 1
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Abstract


Cyclic adenosine monophosphate (cAMP) and cyclic guanosine monophosphate (cGMP) are two important members of the intracellular second messengers. Adenylyl cyclase (AC) and Guanylyl cyclase (GC) to promote the formation of intracellular cAMP and cGMP. And the intracellular concentration of cAMP and cGMP depend on the activity of the phosphodiesterases superfamily Enzymes (PDEs). The PDEs protein family adjust the levels of cAMP and cGMP via catalytic the cGMP and cAMP hydrolysis, intervention in the process of the signal transduction via protein phosphorylation, and the PDEs are also regulated by cGMP and cAMP. At least five of the cyclic nucleotide phosphodiesterase subtypes discovered in the gastric tissue, they are:(1) Ca2+/calmodulin-dependent PDE (PDE1);(2)the cGMP-activated PDE (PDE2);(3)the cGMP-inhibited PDE (PDE3);(4)the cAMP-specific PDE (PDE4);(5)the cGMP-specific PDE (PDE5). C-type natriuretic peptide(CNP) combined with B-type natriuretic peptide receptor(NPR-B) generate cGMP. The CNP via CNP-NPR-B/pGC-cGMP signal transduction pathways inhibition the gastric smooth muscle spontaneous contraction in rats. Therefore, we postulate that NPs may affect the activity of PDEs by adjusting the concentration of intracellular cGMP, regulate the intracellular cAMP concentration, and eventually regulating the gastric antral smooth muscle spontaneous contraction. Based on above premise, we used Forskolin, H-89, CNP, IBMX, EHNA and Milrinone in the isolated muscle strips experiment and enzyme-linked immunosorbent assay (ELISA), explore the activity of the cAMP in gastric antral circular smooth muscle spontaneous contraction, research PDE whether to participate in the CNP pathway of the inhibitory effect of the gastric antral circular smooth muscle spontaneous contraction in rats and related transduction pathway.The dates as follows:1. Different concentrations of Forskolin(10-8mol/L,3×10-8mol/L,10-7mol/L), inhibited the contraction amplitude and lowered the baseline of contraction movement, significantly(n=6,*P<0.05,**P<0.01vsControl.#P<0.05,##P<0.01vs10-8mol/L.&P<0.05vs3×10-8mol/L), dose-dependent. Measured by ELISA, Forskolin concentration-dependently increased the production of cAMP in the perfusate(n=8,*P<0.05,**P<0.01vs Control.#P<0.05,##P<0.01vs10-8mol/L.&P<0.05vs3X10-8mol/L), which showed a significant negative correlation with the contraction amplitude of gastric antral circular muscle(γ=-0.7,P<0.01). The inhibitory effect of Forskolin on spontaneous contraction activity of rat gastric antral circular smooth muscle could be blocked by cAMP-dependent protein kinase (PKA) inhibitor H-89(n=6,*P<0.01).2.(1) Added CNP comparison with the previous, compared with the amplitude/tissue weight, CNP inhibited the contraction amplitude, significantly(n=8,*P<0.01). Measured by ELISA, after added CNP, cGMP content was increased(n=8,*P<0.01), and cAMP content was increased(n=8,**P<0.01), significantly. It is indicated that, CNP probably via generating the cGMP to regulate the PDEs, thereby affecting the level of cAMP.(2) Added CNP in the presence of IBMX compared with only CNP, the inhibition rate of amplitude is decreased, significantly(n=8,*P<0.01). Measured by ELISA, added CNP in the presence of IBMX compared with only CNP, cGMP content is increased(n=8,*P<0.01), significantly. But can’t change the cAMP content(n=8, P>0.05). It is indicated that, IBMX can inhibit the contraction of gastric antral circular smooth muscle, and block part of the CNP-decreased gastric antral smooth muscle spontaneous contraction in rats.(3) Added CNP in the presence of EHNA compared with only CNP, the inhibition rate of amplitude is no difference(n=8, P>0.05). Measured by ELISA, added CNP in the presence of EHNA compared with only CNP, the cGMP and cAMP contents are all further increased(n=8,#P<0.05). It is indicated that, EHNA can inhibit the contraction of gastric antrum circular smooth muscle, but can’t block the CNP-decreased gastric antral smooth muscle spontaneous contraction in rats.(4) Added CNP in the presence of Milrinone compared with only CNP, the inhibition rate of amplitude is decreased, significantly(n=8,*P<0.01). Measured by ELISA, added CNP in the presence of Milrinone compared with only CNP, cGMP content is increased(n=8,#P<0.05), significantly. But can’t change the cAMP content(n=8, P>0.05). It is indicated that, CNP can’t change the cAMP content in the presence of Milrinone, Milrinone participates in the process of the CNP-decreased gastric antral smooth muscle spontaneous contraction in rats.These results suggest that:1. CNP increased intracellular cAMP level via GC-cGMP pathway inhibited PDE3, in the gastric antral smooth muscle cells.2. CNP regulated cAMP content via GC-cGMP-PDE3signaling pathway, and together with cGMP decreased gastric antral smooth muscle spontaneous contraction in rats.

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