Amplification of nitric oxide signaling by interstitial cells isolated from canine colon.

Abstract

The effects of nitric oxide (NO) on intracel- collected by using a video-based imaging system that has been described (23). The locations of cells of interest were specified by using a computer mouse and custom-designed software. Signals were recorded as the fluorescence within a rectangular region over each cell. Fluorescent responses were normalized by dividing by the maximum response to the bath perfusion of a solution containing 1 ,uM Bay K 8644. Thus, the ratio associated with the ordinate of each trace represents a fraction of the increase in [Ca2+]i relative to the response to 1 ,uM Bay K 8644. By continuously framegrabbing images, the computer was able to track the activity of a number of individual cells within a field of view simultaneously. NO was introduced by bolus application of saturated solution (1 mM) to the bath. Bay K 8644 was applied to individual cells by micropressure ejection (2 bars; 15 msec) using a Picospritzer apparatus (General Valve, Fairfield, NJ). Other agents were added or removed from solutions by rapidly exchanging bath medium. MATERIALS AND METHODS CellsHanks' perfusion buffer, NO stock solutions, oxyhemoglobin, and methemoglobin were prepared as described (11, Abbreviations: GI, gastrointestinal; IC, interstitial cell; SMC, smooth muscle cell; [Ca2+]j, intracellular Ca2+ concentration; L-NAME, NG-nitro-L-arginine methyl ester. 2087The publication costs of this article were defrayed in part by page charge payment. This article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. §1734 solely to indicate this fact.

A Fundamental Role for the Nitric Oxide-G-Kinase Signaling Pathway in Mediating Intercellular Ca2+Waves in Glia

“…Because NO synthesis by constitutive nitric oxide synthase (cNOS) is usually calcium-dependent (Moncada et al, 1991), a rise in [Ca 2ϩ ] i may serve to amplify NO production as previously reported (Publicover et al, 1993), and a high rate of potentially unrestricted diffusion for NO could also give rise or contribute to Ca 2ϩ waves seen in many single cells and tissues (Berridge and Dupont, 1994). Therefore, considering the presence of Ca 2ϩ -dependent nitric oxide synthase in glia (Feinstein et al, 1994) and the possibility of cross-talk between NO and Ca 2ϩ in these cells, this study investigated whether the NO-G-kinase signaling pathway is involved in Ca 2ϩ homeostasis in glia and whether a hypothetical mobilization of Ca 2ϩ by NO might initiate regenerative intercellular Ca 2ϩ waves or contribute to intercellular Ca 2ϩ wave propagation induced by mechanical stress of single glial cells in mixed glial-neuron cultures.…”

“…2b,d,e), underscoring a specific effect for NO on intracellular Ca 2ϩ release in glia. It has previously been reported that NO-induced Ca 2ϩ mobilization is ryanodine receptor (RyR)-linked in several cell types (Publicover et al, 1993;Willmott et al, 1995a;Willmott et al, 1996c). To test for the involvement of a RyR-mediated Ca 2ϩ release mechanism, cells were preincubated with an antagonizing concentration of ryanodine before NO application.…”

“…There are however other species that may participate as extracellular messengers in glial intercellular Ca 2ϩ waves. There is increasing evidence that the highly diffusible messenger nitric oxide (NO) can induce Ca 2ϩ mobilization in several cell types (Publicover et al, 1993; Willmott et al, 1995a,b,c;Clementi et al, 1996) either via the cGMP-dependent protein kinase (G-kinase)-coupled activation of ADP-ribosyl cyclase, resulting in an increased synthesis of the potent Ca 2ϩ mobilizing agent cyclic ADP-ribose (Willmott et al, 1996c;Clementi et al, 1996), or via direct nitrosylation of regulatory thiol groups of ryanodine receptors (Stoyanovsky et al, 1997).Because NO synthesis by constitutive nitric oxide synthase (cNOS) is usually calcium-dependent (Moncada et al, 1991), a rise in [Ca 2ϩ ] i may serve to amplify NO production as previously reported (Publicover et al, 1993), and a high rate of potentially unrestricted diffusion for NO could also give rise or contribute to Ca 2ϩ waves seen in many single cells and tissues (Berridge and Dupont, 1994). Therefore, considering the presence of Ca 2ϩ -dependent nitric oxide synthase in glia (Feinstein et al, 1994) and the possibility of cross-talk between NO and Ca 2ϩ in these cells, this study investigated whether the NO-G-kinase signaling pathway is involved in Ca 2ϩ homeostasis in glia and whether a hypothetical mobilization of Ca 2ϩ by NO might initiate regenerative intercellular Ca 2ϩ waves or contribute to intercellular Ca 2ϩ wave propagation induced by mechanical stress of single glial cells in mixed glial-neuron cultures.…”

“…There are however other species that may participate as extracellular messengers in glial intercellular Ca 2ϩ waves. There is increasing evidence that the highly diffusible messenger nitric oxide (NO) can induce Ca 2ϩ mobilization in several cell types (Publicover et al, 1993;Willmott et al, 1995a,b,c;Clementi et al, 1996) either via the cGMP-dependent protein kinase (G-kinase)-coupled activation of ADP-ribosyl cyclase, resulting in an increased synthesis of the potent Ca 2ϩ mobilizing agent cyclic ADP-ribose (Willmott et al, 1996c;Clementi et al, 1996), or via direct nitrosylation of regulatory thiol groups of ryanodine receptors (Stoyanovsky et al, 1997).…”

Ultrastructural features and possible functional role of kit-positive interstitial cells in the guinea pig corpus cavernosum

“…6,[9][10][11] Recent morphological and functional evidence from various organs suggests that ICC and ICs might act as the primary targets for nerve-released NO to have a important role in NO-dependent signal transduction and smooth muscle relaxation. [12][13][14] ICs have been identified by their immunoreactivity to the kit receptor in the corporal tissue from man and several animals. [15][16][17] As dispersed corporal SMCs are able to generate spontaneous electrical activity by opening Ca 2 þ -activated Cl À channels, which is primarily initiated by spontaneous Ca 2 þ store-dependent membrane depolarization, 18,19 the role of ICs in corporal tissues as the primary electrical pacemaker has been questioned; they may serve as intermediaries in the transmission of nerve signals to SMCs or have other functional roles.…”