Effect of entacapone on colon motility and ion transport in a rat model of Parkinson's disease

Abstract

Aim: To study the effects of entacapone, a catechol-O-methyltransferase inhibitor, on colon motility and electrolyte transport in Parkinson's disease (PD) rats.

Methods: Distribution and expression of catechol-O-methyltransferase (COMT) were measured by immunohistochemistry and Western blotting methods. The colonic smooth muscle motility was examined in vitro by means of a muscle motility recording device. The mucosal electrolyte transport of PD rats was examined by using a short-circuit current (ISC ) technique and scanning ion-selective electrode technique (SIET). Intracellular detection of cAMP and cGMP was accomplished by radioimmunoassay testing.

Results: COMT was expressed in the colons of both normal and PD rats, mainly on the apical membranes of villi and crypts in the colon. Compared to normal controls, PD rats expressed less COMT. The COMT inhibitor entacapone inhibited contraction of the PD rat longitudinal muscle in a dose-dependent manner. The β2 adrenoceptor antagonist ICI-118,551 blocked this inhibitory effect by approximately 67% (P < 0.01). Entacapone increased mucosal ISC in the colon of rats with PD. This induction was significantly inhibited by apical application of Cl(-) channel blocker diphenylamine-2, 2'-dicarboxylic acid, basolateral application of Na(+)-K(+)-2Cl(-)co-transporter antagonist bumetanide, elimination of Cl(-) from the extracellular fluid, as well as pretreatment using adenylate cyclase inhibitor MDL12330A. As an inhibitor of prostaglandin synthetase, indomethacin can inhibit entacapone-induced ISC by 45% (P < 0.01). When SIET was applied to measure Cl(-) flux changes, this provided similar results. Entacapone significantly increased intracellular cAMP content in the colonic mucosa, which was greatly inhibited by indomethacin.

Conclusion: COMT expression exists in rat colons. The β2 adrenoceptor is involved in the entacapone-induced inhibition of colon motility. Entacapone induces cAMP-dependent Cl(-) secretion in the PD rat.

Keywords: Catechol-O-methyltransferase; Colon motility; Entacapone; Ion transport; Parkinson disease.

Figure 1

Characteristics of catechol-O-methyltransferase localization in normal and 6-OHDA Parkinson’s disease model rats. A: The COMT immunoreactivity in colon; B: The gray changes of COMT immunoreactivity in colon; C: The changes of expression of COMT in colonic mucosal preparations in 6-OHDA PD rats (n = 8, ^aP < 0.05 vs control). COMT: Catechol-O-methyltransferase; PD: Parkinson’s disease.

Figure 2

Effect of entacapone on colon smooth muscle motility in 6-OHDA Parkinson’s disease rats. A: Inhibitory effect of entacapone on colon longitudinal muscle contraction; B: Dose-response curve effect of entacapone on colon smooth muscle motility in 6-OHDA rats (n = 30); C: Representative tracing of strips when using entacapone (ENT) and β₂ receptor antagonist ICI-118,551 (1.0 × 10^-5 mol/L) in 6-OHDA rats; D: The role of β₂ receptor in entacapone-induced effect of smooth muscle inhibition (n = 9, ^aP < 0.05 vs control).

Figure 3

Entacapone-induced I_SC response in colonic mucosa specimens in 6-OHDA Parkinson’s disease rats. A: The short circuit current (I_SC) changes induced by apical and basolateral using of ENT (200 μmol L^-1); B: Basolateral application of ENT (200 μmol/L) induced transepithelial electric resistance (n = 13); C: Effect of indomethacin (INDO) (10 μmol/L) on the entacapone (ENT)-caused I_SC changes with 6-OHDA PD rats (n = 9); D: Summary of the effects of pretreatment by removing Cl^- from the extracellular fluid, the apical use of DPC (1 mmol/L) (a type of Cl^-channel inhibitor), and the basolateral use of Na⁺-K⁺-2Cl^- co-transporter (NKCC) antagonist bumetanide (100 μmol/L) on the ENT-caused I_SC changes in mucosal preparations of colon in 6-OHDA PD rats (n = 9, ^aP < 0.05, ^bP < 0.01 vs control).

Figure 4

Utilizing scanning ion-selective electrode technique to observe the effect of entacapone on colon mucosa Cl^--flux in 6-OHDA rats. A: Application of entacapone (ENT) (200 μmol/L), diphenylamine-2, 2’-dicarboxylic acid (DPC) (1 mmol/L) and bumetanide (100 μmol/L), causing the typical performance of Cl^—flux in colon specimens in 6-OHDA Parkinson’s disease (PD) rats; B: Generalization of the roles of bumetanide and DPC on the Cl^--flux caused by ENT in colon specimens of 6-OHDA PD rats (n = 10); C: Application of ENT (200 μmol/L), preprocessing by bumetanide or DPC, causing the typical performance of Cl^—flux in colon specimens of 6-OHDA PD rats; D: Generalization of Cl^--flux caused by ENT after treatment by bumetanide and DPC in colonic specimens (n = 9); E: Application of ENT (200 μmol/L), and preprocessing by indomethacin (INDO) (10 μmol/L), causing the typical performance of Cl^—flux in colon specimens in 6-OHDA PD rats; F: Generalization of the roles of INDO on the Cl^--flux caused by ENT (n = 9, ^bP < 0.01 vs control).

Figure 5

cAMP pathway involved in entacapone-induced Cl^-secretion in 6-OHDA Parkinson’s disease rats. A: Representation of effects of MDL 12330A (20 μmol/L) on entacapone (ENT)-induced I_SC changes; B: Generalization of I_SC caused by ENT after treatment by MDL 12330A; C: Generalization of the roles of indomethacin (INDO) on intracellular cAMP content caused by ENT in 6-OHDA Parkinson’s disease (PD) rats; D: Generalization of the roles of INDO on intracellular cGMP content caused by ENT in 6-OHDA PD rats (n = 6,^aP < 0.05 vs control).