Serotonin modifies cytoskeleton and brush-border membrane architecture in human intestinal epithelial cells

Abstract

We previously demonstrated that 5-HT treatment of Caco-2 cells inhibited Na ϩ /H ϩ exchangers (NHE) and Cl Ϫ /OH Ϫ exchange activities via distinct signaling mechanisms. Since regulation of several ion transporters such as NHE3 is influenced by intact cytoskeleton, we hypothesized that 5-HT modifies actin cytoskeleton and/or brush-border membrane architecture via involvement of signaling pathways. Ultrastructural analysis showed that 5-HT (0.1 M, 1 h) treatment of Caco-2 cells caused the apical membrane to assume a convex dome shape that was associated with shortening of microvilli. To examine whether these cellular architecture changes are cytoskeleton driven, we analyzed actin cytoskeleton by fluorescence microscopy. 5-HT induced basal stress fibers with prominent cortical actin filaments via 5-HT3 and 5-HT4 receptor subtypes. This induction was partially attenuated by chelation of intracellular Ca 2ϩ and PKC␣ inhibition (Go6976). In vitro assays revealed that PKC␣ interacted with actin and this association was increased by 5-HT. Our data provide novel evidence that 5-HT-induced signaling via 5-HT3/4 receptor subtypes to cause Ca 2ϩ and PKC␣-dependent regulation of actin cytoskeleton may play an important role in modulation of ion transporters that contribute to pathophysiology of diarrheal conditions associated with elevated levels of 5-HT. microvilli; serotonin and actin; 5-HT and curvature; stress fibers; ion transporter; cytoskeleton SEROTONIN (5-HT) is an important hormone and neurotransmitter that influences diverse functions in the gastrointestinal tract. High levels of 5-HT have been implicated in several diarrheal conditions associated with inflammatory bowel diseases or carcinoid syndrome (3,5,23,37). We have previously shown that 5-HT inhibits Na -dependent PKC␣ (16). In contrast, 5-HT inhibited Cl Ϫ /OH Ϫ exchange activity via a Ca 2ϩ -independent pathway involving Src-kinase induced activation of PKC␦ (31). Furthermore, recent studies have shown that 5-HT causes cytoskeletal reorganization via 5-HT4 receptors to induce pulmonary artery smooth muscle cell migration (10). However, nothing is known regarding the regulation of major cytoskeletal elements such as actin by 5-HT in intestinal epithelial cells.A number of ion transporters such as members of NHE gene family have been shown to interact with the cytoskeleton (40). Microfilaments have been suggested to maintain basal activity of NHE3 and have also been implicated in transducing regulatory signals from receptors or hormones. For example, binding of NHE3 to the cytoskeletal binding proteins is an important factor for its inhibition by cAMP/protein kinase A-, Ca 2ϩ -, or cGMP/PKG-mediated pathways (4,22,24,36,39). Increasing evidence suggests that distinct signaling molecules interact with cytoskeleton to mediate physiological functions (14). In addition, a role for the actin cytoskeleton has emerged during clathrin-and caveolin-mediated endocytosis and exocytosis (29, 34). Our previous studies suggested that 5-HT induced inhibition of NHE activ...

A new host cell internalisation pathway for SadA‐expressing staphylococci triggered by excreted neurochemicals

“…5HT1A activation induces activation of protein kinase C (PKC), Src kinase, and mitogen-activated protein (MAP) kinases and inhibition of AC (Raymond et al, 2001), thus causing a decrease in the cytoplasmic cAMP level (Aune, McGrath, Sarr, Bombara, & Kelley, 1993;Fargin et al, 1989). Both 5HT3 and 5HT4 activation induce F-actin formation by Ca2+-and PKC-dependent pathways (Gill et al, 2008). Therefore, in contrast to 5HT1A, 5HT4 activation increases the intracellular cAMP level (Kuemmerle, Murthy, Grider, Martin, & Makhlouf, 1995) by activating AC (Raymond et al, 2001).…”

“…This finding is explainable because studies by Ataee et al showed that HT-29 cells express various 5HT receptors: 5HT1A, 5HT1B, 5HT3, and 5HT4. Both 5HT3 and 5HT4 activation induce F-actin formation by Ca2+-and PKC-dependent pathways (Gill et al, 2008). 5HT1A activation induces activation of protein kinase C (PKC), Src kinase, and mitogen-activated protein (MAP) kinases and inhibition of AC (Raymond et al, 2001), thus causing a decrease in the cytoplasmic cAMP level (Aune, McGrath, Sarr, Bombara, & Kelley, 1993;Fargin et al, 1989).…”

Regulation of Electroneutral NaCl Absorption by the Small Intestine

“…In Caco-2 cells, 5-HT 4 receptor mediate activation of PKCα, inhibition of Na + /H + exchange activity, and reduction of transcription of NHE3 (129). 5-HT also modifies the brush border architecture which in turn reduces NHE3 function (130). …”

Serotonin regulates amylase secretion and acinar cell damage during murine pancreatitis

“…In both cell systems, 5-HT exerted its regulatory action by modifying and thus activating intracellular proteins, including small GTPases, via a transglutaminase-mediated process known as serotonylation. In addition, recent evidence indicates that serotonylation is found on cytoskeletal proteins including actin and regulates contractility in vascular smooth muscle,16 17 and membrane architecture in intestinal epithelial cells 18. This suggests that 5-HT is also likely to regulate structural processes.…”