Sources and implications of basal nitric oxide in spontaneous contractions of guinea pig taenia caeci

Catalina Caballero-Alomar,1 Carmen Santos,1 Diego Lopez,2 M. Teresa Mitjavila,2 and Pere Puig-Parellada1

1Unitat de Farmacologia, Facultat de Medicina, Universitat de Barcelona, 08036 Barcelona; and 2Departament de Fisiologia, Facultat de Biologia, Universitat de Barcelona, E-08028 Barcelona, Spain

Submitted 8 July 2002 ; accepted in final form 9 June 2003


    ABSTRACT
 TOP
 ABSTRACT
 MATERIALS AND METHODS
 RESULTS
 DISCUSSION
 DISCLOSURES
 REFERENCES
 
We examined in vitro the source and role of basal nitric oxide (NO) in proximal segments of guinea pig taenia caeci in nonadrenergic, noncholinergic (NANC) conditions. Using electron paramagnetic resonance (EPR), we measured the effect of the NO synthase inhibitor NG-nitro-L-arginine methyl ester (L-NAME, 104 M), the neuronal blocker tetrodotoxin (TTX, 106 M), or both on spontaneous contractions and on the production of basal NO. Both L-NAME and TTX, when tested alone, increased the amplitude and frequency of contractions. NO production was abolished by L-NAME and was inhibited by 38% by TTX. When tested together, L-NAME in the presence of TTX or TTX in the presence of L-NAME had no further effect on the amplitude or frequency of spontaneous contractions, and the NO production was inhibited. These findings suggest that basal NO consists of TTX-sensitive and TTX-resistant components. The TTX-sensitive NO has an inhibitory effect on spontaneous contractions; the role of TTX-resistant NO is unknown.

longitudinal smooth muscle; cecum; nonadrenergic, noncholinergic innervation; tetrodotoxin; electron paramagnetic resonance


NONADRENERGIC, NONCHOLINERGIC (NANC) inhibitory innervation was first demonstrated in guinea pig taenia caeci (6). Nitric oxide (NO) is a mediator of NANC relaxation after neuronal stimulation in a wide range of intestinal smooth muscle preparations. The presence of NO in relaxation induced by electrical stimulation has been reported in guinea pig taenia caeci (7, 20, 36).

The membrane potential of smooth muscle cells from the gastrointestinal tract usually displays slow rhythmic waves that originate cyclic contractions in many gastrointestinal smooth muscles. Interstitial cells of Cajal (ICCs) were first described in 1893 (8), and Thuneberg in 1982 (37) suggested that they were probably pacemaker cells. In addition to the generation of slow waves, ICCs have recently been shown to act as intermediates in neurotransmission to smooth muscle in the gastrointestinal tract (41, 44).

A number of studies (3, 18, 19, 22, 30, 42) suggest that the contractile activity of various gastrointestinal muscles can be suppressed by basal NO, because spontaneous contractions or tone are enhanced by inhibiting NO synthases (NOS) or by using oxyhemoglobin as scavenger for NO. The opposite effect is obtained by using NO (17, 28) or donors of NO (19). The source of basal NO can be neuronal (5, 19, 43, 45) or extraneuronal (11, 13, 32, 33, 44, 46). However, the presence and role of NO in spontaneous contractions has not been studied in guinea pig taenia caeci so far. This fact, together with the results we obtained (17) showing that the electrical release of NO is inversely proportional to frequency, prompted us to study the role and origin of NO in guinea pig taenia caeci in basal conditions, in which our preparations display rhythmic contractions.

In physiological conditions, NO from constitutive NOS plays an important homeostatic role in the regulation of both secretion and motility in the gastrointestinal tract. Studies on phenotypes of neuronal NOS (nNOS) knockout mice have shown the enlargement of the stomach and pyloric stenosis (14). Furthermore, a decrease in or absence of NO has been implicated in human gastrointestinal disorders (4, 23, 38, 39).

The purpose of the present study was to investigate the role and the source of basal NO in spontaneous contractions of proximal segments of guinea pig taenia caeci.


    MATERIALS AND METHODS
 TOP
 ABSTRACT
 MATERIALS AND METHODS
 RESULTS
 DISCUSSION
 DISCLOSURES
 REFERENCES
 
Preparations of proximal segments of guinea pig taenia caeci and isometric tension recording. Male albino guinea pigs weighing 300–350 g were fasted, with free access to water, for 24 h before they were killed by a blow to the head followed by exsanguination. These procedures were approved by the University of Barcelona Ethical Committee for Animal Experimentation. The cecum was exposed via midline incision. Silk ligatures were tied to each end of dorsal and ventral taenia: the end near the colon was defined as the distal segment and the opposite end as the proximal segment. The taenia was dissected out and placed in Krebs solution (composition in mM: 119.78 NaCl, 4.7 KCl, 7 MgSO4, 1.17 H2O, 1.17 KH2PO4, 24.9 NaHCO3, 11.1 glucose, and 2.56 CaCl2 · 2H2O). Atropine (106 M) and guanethidine (4 x 106 M) were routinely added to the Krebs solution to perform the experiments in NANC conditions. Segments of 1–2 cm were cut from the proximal part of each taenia and were mounted in thermostatically controlled (37 ± 0.5°C) organ baths containing Krebs solution. The pH of the organ bath fluid was kept constant at 7.4 by bubbling the Krebs solution with 95% O2-5% CO2. A resting tension of 2 g was applied to each strip, and the responses were recorded by an isometric force transducer (Harvard VF-1) connected to an amplifier and to a polygraph chart recorder or a computer, in which case data were digitized and simultaneously displayed and collected by using Datawin 1 software (Panlab, Barcelona, Spain) coupled to an ISC-16A/D card.

Experimental protocols. During an equilibration period of 30–60 min, the preparations gradually relaxed to a continuous baseline tension (<0.5 g) and showed spontaneous contractions, the amplitude of which are expressed in millinewtons; the time interval between the peak contractions is expressed in seconds, and the frequency of contractions is expressed in contractions per minute (cpm). Data are compared for statistical purposes as a mean amplitude and frequency during 3–5 contractions before and after adding drugs. When sodium nitroprusside (SNP) was added, we measured the time interval between the peak amplitude of contractions before and after the addition of SNP. Data are expressed as a percentage of the values obtained in the absence of SNP (control). Only 1 or 2 doses of SNP were tested for each preparation to avoid tolerance. When NG-nitro-L-arginine methyl ester (L-NAME) was added in the presence of tetrodotoxin (TTX) or TTX was added in the presence of L-NAME, we waited until the effect of the first drug stabilized before adding the second.

Electrical field stimulation. Some segments with spontaneous contractions were stimulated between parallel platinum electrodes (0.3–1 Hz, 40 V, 1 ms), and the stimulus was applied at the peak of spontaneous contraction for 5 s (7). Electrical field stimulation produced a transient relaxation followed by a frequency-dependent contraction after the stimulation was stopped. TTX (106 M) inhibited electrical relaxation.

Basal NO production by electron paramagnetic resonance. NO production was measured by electron paramagnetic resonance (EPR) (2, 16, 25, 26). Proximal taenia caeci segments were preincubated at 37°C for 15 min in the Krebs solution containing atropine (106 M) and guanethidine (4 x 106 M). They were then exposed to the spin-trapping agents diethyldithiocarbamic acid (DETC) (5 x 103 M) and FeSO4 · 7H2O (5 x 105 M) and were incubated for 30 min. Segments were then weighed and frozen in liquid N2 for posterior EPR analysis. Some segments were preincubated with L-NAME (104 M), TTX (106 M), or both, in this case with an interval of 2 min between the addition of drugs. The mononitrosyl iron complex formed with DETC was measured in a Bruker 300E spectrometer (Bruker Instruments, Billerica, MA) at 77 K (10-mW microwave power, 31,985-G amplitude modulation, 9.77-kHz microwave frequency, and 100-kHz modulation frequency). The results are expressed as intensity units/mg wet tissue. A standard curve of Fe-DETC-NO was generated by diethylamine NONOate (DEA/NO, from 109 to 106 M). This curve was used to extrapolate the Fe-DETC-NO signal and also to estimate the linearity of the assay. The signal is linear (y = 12635x + 108448) in the interval of DEA/NO concentrations between 5 nM (500 units) and 500 nM (30,000 units).

Drugs. The following drugs were used: atropine sulfate, guanethidine sulfate, L-NAME, L-arginine hydrochloride, D-arginine hydrochloride, SNP, TTX, FeSO4 · 7H2O, and DETC, all from Sigma-Aldrich (Madrid, Spain), and DEA/NO, from Cayman Chemical (Ann Arbor, MI).

Statistical analysis. Results were subjected to paired or unpaired analysis of variance, and differences were tested by the Student-Newman-Keuls test (n = number of animals). Data are given as means ± SE. Differences were considered significant when P < 0.05.


    RESULTS
 TOP
 ABSTRACT
 MATERIALS AND METHODS
 RESULTS
 DISCUSSION
 DISCLOSURES
 REFERENCES
 
Spontaneous contractile activity. Spontaneous contractions were observed (Fig. 1) with an amplitude of 23.7 ± 1.2 mN and a time interval between the peak amplitude of 265.2 ± 17.2 s, which corresponds to a frequency of 0.36 ± 0.02 cpm (pooled data n >= 29 animals).



View larger version (33K):
[in this window]
[in a new window]
 
Fig. 1. Effect of tetrodotoxin (TTX, 106 M at arrow) and NG-nitro-L-arginine methyl ester (L-NAME, 104 M at arrow) in the presence of TTX on spontaneous contractions of isolated proximal segment of guinea pig taenia caeci.

 

The basal NO production assessed by EPR is shown in Fig. 2. In the control group, a value of 1,019 ± 39.7 intensity units/mg wet tissue was obtained.



View larger version (13K):
[in this window]
[in a new window]
 
Fig. 2. Inhibitory effect of L-NAME (104 M), TTX (106 M), or both on basal nitric oxide (NO) production in isolated proximal segments of guinea pig taenia caeci. Data are given as means ± SE of intensity units of electron paramagnetic resonance (EPR)/mg wet tissue for 3–7 animals. *P < 0.05 vs. control.

 

NO inhibition. To determine the role of basal NO in spontaneous contractions, we added L-NAME (104 M) to the bath. The frequency and amplitude of contractions were significantly increased [from 0.32 ± 0.04 to 0.48 ± 0.06 cpm (P < 0.001) and from 22.1 ± 1.9 to 26.7 ± 2.2 mN (P < 0.01); n = 12], whereas the basal tone was not affected (Fig. 3). In the presence of L-arginine (103 M), the effects of L-NAME on the frequency and the amplitude were abolished (control, 0.33 ± 0.03 and L-arginine + L-NAME, 0.31 ± 0.03 cpm; control, 26.4 ± 2.5 and L-arginine + L-NAME, 26.3 ± 3.2 mN; n = 4) (Fig. 4) but D-arginine (103 M) had no effect [control, 0.31 ± 0.05 and D-arginine + L-NAME, 0.47 ± 0.06 cpm (P < 0.01); control, 19.4 ± 2.6 and D-arginine + L-NAME, 22.6 ± 2.5 mN (P < 0.05); n = 3]. The basal production of NO as determined by EPR was inhibited by 97% by L-NAME when compared with the control group (L-NAME, 31.8 ± 6.5 vs. control, 1,026 ± 34.5 intensity units/mg wet tissue; P < 0.001; n = 3; Fig. 2).



View larger version (15K):
[in this window]
[in a new window]
 
Fig. 3. Effect of TTX (106 M) on the frequency and amplitude of spontaneous contractions of isolated proximal segments of guinea pig taenia caeci before and after L-NAME (104 M). Data are expressed as means ± SE for 10–12 animals and are reported as %control (with control = 100%). *P < 0.05 vs. control by paired analysis of variance.

 


View larger version (20K):
[in this window]
[in a new window]
 
Fig. 4. A representative register of the effect of L-NAME 104 M in the presence of 103 M L-arginine (L-Arg) on spontaneous contractions of isolated proximal segment of guinea pig taenia caeci.

 

Responses to SNP. The NO donor SNP produced a dose-dependent prolongation of the interval between spontaneous contractions immediately before and after the addition of SNP at 108 (237.6 ± 69.9 to 232.7 ± 66.8 s; n = 4), at 107 (308.5 ± 49.2 to 535.9 ± 120.7 s; n = 10; P < 0.05), and at 106 M (208.8 ± 33.0 to 433.3 ± 95.8 s; n = 10; P < 0.001) (Fig. 5). When the effect of SNP decayed and the spontaneous contractions restarted, we observed a decrease in amplitude and frequency of spontaneous contraction with respect to control.



View larger version (20K):
[in this window]
[in a new window]
 
Fig. 5. Inhibitory effect of sodium nitroprusside (SNP) on spontaneous contractions of isolated proximal segment of guinea pig taenia caeci. A: representative effect of SNP (106 M) on spontaneous contractions. B: effect of SNP (108, 107, 106 M) on the interval between spontaneous contractions immediately before and after the addition of SNP in isolated proximal segments of guinea pig taenia caeci. Data are expressed as means ± SE for 4–10 animals and are reported as %control. *P < 0.05 vs. control. ***P < 0.001 vs. control.

 

We checked that SNP generated Fe-DETC-NO in vitro when incubated with Fe2+ and DETC. A concentration of 108 M produced a slight signal. The signals corresponding to 107 and 106 M of SNP produced 800 and 9,000 nmol/10 ml · 15 min, respectively.

Effect of TTX or neuronal NO. Electrical field stimulation applied at the peak of spontaneous contractions produced a relaxation (39.0 ± 3.3 and 90.2 ± 3.7% at 0.3 and 1 Hz, 40 V, 1 ms, respectively; n = 3) that was abolished by TTX (106 M) (Fig. 6). TTX on spontaneous contractions significantly increased the amplitude (from 26.6 ± 3.0 to 29.0 ± 3.1 mN; n = 10; P < 0.01) and frequency (from 0.27 ± 0.03 to 0.40 ± 0.05 cpm; n = 10; P < 0.01) (Fig. 1). Basal production of NO in the presence of TTX was 62% of control (TTX, 633.3 ± 38.8 vs. control, 1,019.25 ± 39.7 intensity units/mg wet tissue; n = 7; P < 0.0001) (Fig. 2).



View larger version (27K):
[in this window]
[in a new window]
 
Fig. 6. Inhibitory effect of TTX (106 M) on nonadrenergic, noncholinergic relaxation induced by electrical field stimulation (EFS) on a proximal segment of guinea pig taenia caeci. Stimulation at 0.6 Hz, 40 V, 1 ms was applied for 5 s at the peak of the spontaneous contractions.

 

Effect of TTX + L-NAME or additional NO. The effect of TTX on spontaneous movements was similar to that of L-NAME (104 M) (Figs. 1 and 3), although the production of NO was partially blocked by TTX (38%), in contrast to L-NAME, which blocked it totally. According to these results, we deduced that TTX-sensitive NO or neuronal NO is the regulator of spontaneous movements. To confirm this and to check the role of nonneuronal NO in spontaneous movements, we added L-NAME (total NO blockade) after TTX (neuronal blockade). In these conditions, if TTX-resistant NO had a role, frequency and/or amplitude should be affected as occurred in canine (19) and rat colon amplitude (24) and in canine lower esophageal sphincter (10). However, no significant differences were observed in either frequency (TTX, 0.40 ± 0.05 vs. TTX + L-NAME, 0.41 ± 0.05 cpm) or amplitude (TTX, 29.0 ± 3.1 vs. TTX + L-NAME, 27.9 ± 3.2 mN; n = 10) of contractions (Fig. 1). We thus discarded the possibility that extraneuronal NO had an important role in the regulation of spontaneous movements. These results also confirm the relevant role of neuronal NO according to the findings of other authors (19, 24, 27). The basal production of NO in this condition was abolished [TTX, 633.3 ± 38.8 (n = 7) and TTX + L-NAME, 37.5 ± 7.1 (n = 3) intensity units/mg wet tissue; Fig. 2].

Effect of L-NAME + TTX or additional neurotransmitters. In basal conditions, we have demonstrated that neuronal NO release is produced, but we cannot rule out the release of other neurotransmitters. Like other authors (19, 24, 27), to assess this possibility, we added TTX (neuronal blockade) after L-NAME (total inhibition of NO). In these conditions, if neurotransmitters other than NO were released, the frequency and/or amplitude of spontaneous contractions should be affected. However, no significant differences were observed in frequency, although it decreased slightly (from 0.48 ± 0.06 to 0.40 ± 0.07 cpm; n = 12). Nor were differences observed in the amplitude (L-NAME, 26.7 ± 2.1 vs. L-NAME + TTX, 26 ± 1.9 mN; n = 12) of contractions (Fig. 3), which allowed us to rule out the action of other neurotransmitters, and again these results confirm the relevant role of neuronal NO. The basal production of NO in the presence of L-NAME + TTX was abolished (control, 1,026 ± 34.5 vs. L-NAME + TTX, 31.6 ± 5.8 intensity units/mg wet tissue; n = 3) as obtained with L-NAME alone (31.7 ± 6.5 intensity units/mg wet tissue; n = 4) or TTX + L-NAME (37.5 ± 7.1 intensity units/mg wet tissue; n = 3) (Fig. 2).


    DISCUSSION
 TOP
 ABSTRACT
 MATERIALS AND METHODS
 RESULTS
 DISCUSSION
 DISCLOSURES
 REFERENCES
 
We examined the relation between the release of basal NO and spontaneous contractions of guinea pig taenia caeci. Basal NO in NANC conditions has two components: TTX-sensitive NO and TTX-resistant NO. The TTX-sensitive NO or neuronal NO inhibits spontaneous contractions.

Studies on other parts of the intestine and other animal species have implicated NO in spontaneous contractions (9, 19, 22, 24, 27, 31). L-NAME, an inhibitor of NOS, abolished the production of NO and increased the frequency and amplitude of spontaneous contractions in taenia caeci. These observations suggested that the basal NO is produced continuously and in a sufficiently high concentration to maintain inhibition of these parameters.

NO relaxes the smooth muscle by causing an increase in cGMP, which in turn mediates smooth muscle relaxation (35). Our data show that SNP produced a dose-dependent inhibition of spontaneous contractions that can be explained by the results of Kwon et al. (21), who demonstrated that SNP, through cGMP-dependent signaling pathways, inhibits myosin light chain phosphorylation and smooth muscle contraction by decreasing intracellular Ca2+ concentration in guinea pig taenia caeci. However, SNP yields several forms of redox NO that produce smooth muscle hyperpolarization by various ionic mechanisms (12). Thus our results may not only reflect the response to endogenous NO.

The production of basal NO in the presence of TTX was 62% of the total, and so the TTX-sensitive NO was 38%. TTX completely abolished the relaxation induced by electrical stimulation. On spontaneous movements, TTX increased the frequency and amplitude of spontaneous contractions, in agreement with studies on other tissues (19, 24, 31) and confirming that the pacemaker activity is not neurogenic.

We have demonstrated by EPR that the NO not blocked by TTX is released in nearly twice the amount of the neuronal NO (Fig. 2). If this NO had a modulator role in spontaneous contraction like the neuronal NO, L-NAME after TTX should produce significant variations in spontaneous contractions. Our results do not show significant differences either in frequency or in amplitude, despite the large amount of TTX-resistant NO released. Consistent with this finding, no changes in amplitude in rat proximal colon were observed by other authors (27). These results confirm the relevant role of neuronal NO and indicated that the additional NO had different functions and mechanisms from neuronal NO. In canine lower esophageal sphincter, myogenic NOS activity has been observed (33) and a different mechanism of action for myogenic NO and neurogenic NO has been proposed (10). Both neuronal and myogenic NO induced relaxation; thus when NOS activity was blocked by NG-nitro-L-arginine (L-NNA), a NOS inhibitor, after enteric nerve activities were inhibited by TTX, tone persistently increased. In similar experimental conditions, an increase in the force or amplitude of contractions has been observed in distal colonic circular smooth muscle of rat (24) and canine colon (19), and the authors suggested an additional, extraneuronal or potential-independent release of NO. Martinez-Cuesta et al. (22) reported that the contractile response of the rat isolated duodenum to L-NNA was greatly inhibited in the presence of TTX, which was attributed to neuronal involvement of NO. However, the present paper shows that the basal production of NO in taenia caeci measured by EPR is similar with L-NAME or TTX + L-NAME, which means that TTX does not block the effect of L-NAME. On the other hand, the discrepancies observed among the tissues assayed may be due to the amount of NO released and their possible action(s). All of these effects are likely to depend on the species, tissue, and region considered.

The enzymes that generate NO, known as NOS, are encoded by three genes located on separate chromosomes. The three NOS isoforms known are type I or nNOS, type II or inducible NOS (iNOS), and type III or endothelial NOS (eNOS). NOS has been identified in the myenteric plexus and submucous plexus from the esophagus to the rectum in guinea pig (29), and it could be the origin of TTX-sensitive or neuronal NO released in our basal conditions, although we cannot rule out the presence of eNOS and iNOS (40). The source of the 62% of TTX-resistant NO could be the eNOS, nNOS, or iNOS released by the interstitial cells (40, 44, 46), smooth muscle (10, 13, 15, 33), or neurons with TTX-resistant sodium-channel currents (1); action potential-independent release of NO from neurons; and finally the NO generated by the mitochondrial NOS that exerts a substantial control over mitochondrial respiration (11).

The primary target of TTX-sensitive basal NO is unknown. NO has multiple actions in postjunctional cells of the gastrointestinal tract. Several theories of NO-dependent muscle relaxation and the role of ICCs have been discussed (34, 35). Our results implicate TTX-sensitive basal NO in the frequency and amplitude of spontaneous contractions. It is more difficult to identify the role and the target of TTX-resistant basal NO, but the fact that L-NAME in the presence of TTX does not produce changes suggests that the TTX-resistant NO may be implicated in different roles of TTX-sensitive basal NO.

L-NNA as L-NAME produced incomplete inhibition of electrically induced relaxation on taenia caeci (7); thus other neurotransmitters are probably released, together with NO. TTX completely blocked the electrically induced relaxation. In basal conditions we have demonstrated that neuronal NO release is produced, but we cannot rule out the release of other neurotransmitters. To assess this possibility, we added L-NAME and observed an increase of frequency and amplitude of contractions (attributed to NO block), and after adding the TTX there was no alteration in amplitude or frequency, discarding the role of other neurotransmitters. These observations are in agreement with other studies in rat distal colon (24), in rat proximal colon (27), and in canine colon (19). Thus neuronal NO may be the only neurotransmitter released in basal NANC conditions involved in the control of spontaneous contractions.

In conclusion, our results demonstrate that in NANC basal conditions the proximal segments of taenia caeci continuously release NO. Basal NO has two components, one that is TTX sensitive and another that is TTX resistant. TTX-sensitive NO mediates the inhibition of spontaneous movements. The role of TTX-resistant NO is unknown. These results may help us to understand mechanisms of gut motility.


    DISCLOSURES
 TOP
 ABSTRACT
 MATERIALS AND METHODS
 RESULTS
 DISCUSSION
 DISCLOSURES
 REFERENCES
 
C. Santos was funded by a fellowship awarded by the Agustí Pedro i Pons Foundation (University of Barcelona).


    ACKNOWLEDGMENTS
 
We thank Professor J. Forn for useful advice and Robin Rycroft of the Language Advisory Service of the University of Barcelona for help in editing the manuscript.


    FOOTNOTES
 

Address for reprint requests and other correspondence: C. Caballero-Alomar, Unitat de Farmacologia, Facultat de Medicina, Universitat de Barcelona, Casanova 143, 08036 Barcelona, Spain (E-mail catalina{at}medicina.ub.es).

The costs of publication of this article were defrayed in part by the payment of page charges. The article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.


    REFERENCES
 TOP
 ABSTRACT
 MATERIALS AND METHODS
 RESULTS
 DISCUSSION
 DISCLOSURES
 REFERENCES
 

  1. Akopian AN, Sivilotti L, and Wood JN. A tetrodotoxin-resistant voltage-gated sodium channel expressed by sensory neurons. Nature 379: 257–262, 1996.[ISI][Medline]
  2. Benito S, Lopez D, Sáiz MP, Buxaderas S, Sánchez J, Puig-Parellada P, and Mitjavila MT. A flavonoid-rich diet increases nitric oxide production in rat aorta. Br J Pharmacol 135: 910–916, 2002.[Abstract/Free Full Text]
  3. Boeckxstaens GE, Pelckmans PA, Bult H, De Man JG, Herman AG, and Van Maercke YM. Non-adrenergic noncholinergic relaxation mediated by nitric oxide in the canine ileocolonic junction. Eur J Pharmacol 190: 239–246, 1990.[ISI][Medline]
  4. Boughton-Smith NK, Evans SM, Hawkey CJ, Cole AT, Balsitis M, Whittle BJR, and Moncada S. Nitric oxide synthase activity in ulcerative colitis and Crohn's disease. Lancet 342: 338–340, 1993.[ISI][Medline]
  5. Bredt DS, Hwang PM, and Snyder SH. Localization of nitric oxide synthase indicating a neuronal role for nitric oxide. Nature 347: 768–770, 1991.[ISI]
  6. Burnstock G, Campbell G, and Rand MJ. The inhibitory innervation of the taenia of the guinea-pig caecum. J Physiol 182: 504–526, 1966.[ISI][Medline]
  7. Caballero-Alomar C, Santos C, and Puig-Parellada P. Evidence that inhibitory neurotransmission differs between the proximal and distal segments of guinea-pig taenia caeci. Eur J Pharmacol 369: 215–219, 1999.[ISI][Medline]
  8. Cajal SR. Sur les ganglions et plexus nerveux de l'intestin. C R Soc Biol (Paris) 45: 217–223, 1893.
  9. Calignano A, Whittle BJR, Di Rosa M, and Moncada S. Involvement of endogenous nitric oxide in the regulation of rat intestinal motility in vivo. Eur J Pharmacol 229: 273–276, 1992.[ISI][Medline]
  10. Daniel EE, Jury J, Salapateck AM, Bowes T, Lam A, Thomas S, Ramnarain M, Nguyen V, and Mistry V. Nitric oxide from enteric nerves acts by a different mechanism from myogenic nitric oxide in canine lower esophageal sphincter. J Pharmacol Exp Ther 294: 270–279, 2000.[Abstract/Free Full Text]
  11. Ghafourifar P and Richter C. Nitric oxide synthase activity in mitochondria. FEBS Lett 418: 291–296, 1997.[ISI][Medline]
  12. Goyal RK and Xue DH. Evidence for NO · redox form of nitric oxide as nitrergic inhibitory neurotransmitter in gut. Am J Physiol Gastrointest Liver Physiol 275: G1185–G1192, 1998.[Abstract/Free Full Text]
  13. Grider JR, Murthy KS, Jin JG, and Makhlouf GM. Stimulation of nitric oxide from muscle cells by VIP: prejunctional enhancement of VIP release. Am J Physiol Gastrointest Liver Physiol 262: G774–G778, 1992.[Abstract/Free Full Text]
  14. Huang PL. Molecular biology of nitric oxide synthase genes and nitric oxide synthase-knockout mice. In: Nitric Oxide and the Peripheral Nervous System, edited by Toda N, Moncada S, Furchogott R, and Higgs EA. London: Portland, 2000, p. 17–35.
  15. Jin JG, Murthy J, Grider JR, and Makhlouf GM. Stoichiometry of neurally induced VIP release, NO formation, and relaxation in rabbit and rat gastric muscle. Am J Physiol Gastrointest Liver Physiol 271: G357–G369, 1996.[Abstract/Free Full Text]
  16. Kalyanaraman B. Detection of nitric oxide by electron spin resonance in chemical, photochemical, cellular, physiological, and pathophysiological systems. Methods Enzymol 268: 168–187, 1996.[ISI][Medline]
  17. Kanada A, Hata F, Suthamnatpong N, Machara T, Ishii T, and Yagasaki O. Key roles of nitric oxide and cyclic GMP in nonadrenergic and noncholinergic inhibition in rat ileum. Eur J Pharmacol 216: 287–292, 1992.[ISI][Medline]
  18. Keef KD, Du C, Ward SM, McGreggor B, and Sanders KM. Enteric inhibitory neuronal regulation of human colonic circular muscle: role of nitric oxide. Gastroenterology 105: 1009–1016, 1993.[ISI][Medline]
  19. Keef KD, Murray DC, Sanders KM, and Smith TK. Basal release of nitric oxide induces an oscillatory motor pattern in canine colon. J Physiol 4993: 773–786, 1997.
  20. Knudsen MA and Tottrup A. A possible role of the L-argininenitric oxide pathway in the modulation of cholinergic transmission in the guinea-pig taenia coli. Br J Pharmacol 107: 837–841, 1992.[Abstract]
  21. Kwon SC, Ozaki H, and Karaki H. NO donor sodium nitroprusside inhibits excitation-contraction coupling in guinea pig taenia coli. Am J Physiol Gastrointest Liver Physiol 279: G1235–G1241, 2000.[Abstract/Free Full Text]
  22. Martinez-Cuesta MA, Esplugues JV, and Whittle BJR. Modulation by nitric oxide of spontaneous motility of the rat isolated duodenum: role of tachynins. Br J Pharmacol 118: 1335–1340, 1996.[Abstract]
  23. Mearin F, Mourelle M, Guarner F, Salas A, RiverosMoreno VS, and Malagelada JR. Patients with achalasia lack nitric oxide synthase in the gastro-oesophageal junction. Eur J Clin Invest 23: 724–728, 1993.[ISI][Medline]
  24. Middleton SJ, Cuthbert AW, Shorthouse M, and Hunter JO. Nitric oxide affects mammalian distal colonic smooth muscle by tonic neuronal inhibition. Br J Pharmacol 108: 974–979, 1993.[Abstract]
  25. Mikoyan VD, Kubrina LN, Serezhenkov VA, Stukan RA, and Vanin AF. Complexes of Fe2+ with diethyldithiocarbamate or N-methyl-D-glucamine dithiocarbamate as traps of nitric oxide in animal tissues: comparative investigations. Biochim Biophys Acta 1336: 225–234, 1997.[ISI][Medline]
  26. Mordvintcev P, Mulsch A, Busse R, and Vanin AF. On-line detection of nitric oxide formation in liquid aqueous phase by electron paramagnetic resonance spectroscopy. Anal Biochem 199: 142–146, 1991.[ISI][Medline]
  27. Mulé FD, D'Angelo S, Amato A, Contino I, and Serio R. Modulation by nitric oxide of spontaneous mechanical activity in rat proximal colon. J Auton Pharmacol 19: 1–6, 1999.[ISI][Medline]
  28. Murr MM, Balsiger BM, Farrugia G, and Sarr MG. Role of nitric oxide, vasoactive intestinal polypeptide, and ATP in inhibitory neurotransmission in human jejunum. J Surg Res 84: 8–12, 1999.[ISI][Medline]
  29. Nichols K, Krantis A, and Staines W. Histochemical localization of nitric oxide-synthesizing neurons and vascular sites in the guinea-pig intestine. Neuroscience 51: 791–799, 1992.[ISI][Medline]
  30. Ozaki H, Blondfield DP, Hori M, Publicover NG, Kato I, and Sanders KM. Spontaneous release of nitric oxide inhibits electrical, Ca2+ and mechanical transients in canine gastric smooth muscle. J Physiol 445: 231–237, 1992.[Abstract]
  31. Plujà L, Fernández E, and Jiménez M. Neuronal modulation of the cyclic electrical and mechanical activity in the rat colonic circular muscle: putative role of ATP and NO. Br J Pharmacol 126: 883–892. 1999.[Abstract/Free Full Text]
  32. Pollock JS, Nakane M, Buttery LD, Martinez A, Springall D, Polak JM, Fostermann U, and Murad F. Characterization and localization of endothelial nitric oxide synthase using specific monoclonal antibodies. Am J Physiol Cell Physiol 265: C1379–C1387, 1993.[Abstract/Free Full Text]
  33. Salapatek AM, Wang YF, Mao YK, Lam A, and Daniel EE. Myogenic nitric oxide synthase activity in canine lower oesophageal sphincter: morphological and functional evidence. Br J Pharmcol 123: 1055–1064, 1998.[Abstract]
  34. Salmhofer H, Neuhuber WL, Ruth P, Huber A, Russwurm M, and Allescher HD. Pivotal role of interstitial cells of Cajal in the nitric oxide signaling pathway of rat small intestine. Cell Tissue Res 305: 331–340, 2001.[ISI][Medline]
  35. Sanders KM and Keef KD. The role of nitric oxide as a neurotransmitter in the gastrointestinal tract. In: Nitric Oxide and the Peripheral Nervous System, edited by Toda N, Moncada S, Furchgott R, and Higgs EA. London: Portland, 2000, p. 115–130.
  36. Shuttleworth CWR, Sweeney KM, and Sanders KM. Evidence that nitric oxide acts as an inhibitory neurotransmitter supplying tenia from guinea-pig caecum. Br J Pharmacol 127: 1495–1501, 1999.[Abstract/Free Full Text]
  37. Thuneberg L. Interstitial cells of Cajal. Intestinal pacemakers cells. Adv Anat Embryol Cell Biol 71: 1–130, 1982.[ISI][Medline]
  38. Vanderwinden JM, De Laet MH, Schiffmann SN, Mailleux P, Lowenstein CJ, Snyder SH, and Vanderhaeghen JJ. Nitric oxide synthase distribution in the enteric nervous system of Hirschsprung's disease. Gastroenterology 104: 969–973, 1993.
  39. Vanderwinden JM, Mailleux P, Schiffmann SN, Vanderhaeghen JJ, and De Laet MH. Nitric oxide synthase activity in infantile hypertrophic pyloric stenosis. N Engl J Med 327: 511–515, 1992.[Abstract]
  40. Vannucchi MG, Corsani L, Bani D, and Faussone-Pellegrini MS. Myenteric neurons and interstitial cells of Cajal of mouse colon express several nitric oxide synthase isoforms. Neurosci Lett 326: 191–195, 2002.[ISI][Medline]
  41. Ward SM, Beckett EAH, Wang XY, Baker F, Khoyi M, and Sanders KM. Interstitial cells of Cajal mediate cholinergic neurotransmission from enteric motor neurons. J Neurosci 20: 1393–1403, 2000.[Abstract/Free Full Text]
  42. Ward SM, Dalziel HH, Bradley ME, Buxton ILO, Keef KD, Westfall DP, and Sanders KM. Involvement of cyclic GMP in non-adrenergic, non-cholinergic inhibitory neurotransmission in dog proximal colon. Br J Pharmacol 107: 1075–1082, 1992.[Abstract]
  43. Ward SM, Dalziel HH, Khoyi MA, Westfall A, Sanders KM, and Westfall DP. Hyperpolarization and inhibition of contraction mediated by nitric oxide released from enteric inhibitory neurones in guinea-pig taenia coli. Br J Pharmacol 118: 49–56, 1996.[Abstract]
  44. Ward SM, Morris G, Reese L, Wang XY, and Sanders KM. Interstitial cells of Cajal mediate enteric inhibitory neurotransmission in the lower esophageal and pyloric sphincters. Gastroenterology 115: 314–329, 1998.[ISI][Medline]
  45. Ward SM, Xue C, Shuttleworth CW, Bredt DS, and Snyder KM. NADPH diaphorase and nitric oxide synthase colocalization in enteric neurons of canine proximal colon. Am J Physiol Gastrointest Liver Physiol 263: G277–G284, 1992.[Abstract/Free Full Text]
  46. Xue C, Pollock J, Schmidt HHW, Ward SM, and Sanders KM. Expression of nitric oxide synthase by interstitial cells of the canine proximal colon. J Auton Nerv Syst 49: 1–14, 1994.[ISI][Medline]




This Article
Abstract
Full Text (PDF)
All Versions of this Article:
285/4/G747    most recent
00273.2002v1
Alert me when this article is cited
Alert me if a correction is posted
Citation Map
Services
Email this article to a friend
Similar articles in this journal
Similar articles in ISI Web of Science
Similar articles in PubMed
Alert me to new issues of the journal
Download to citation manager
Search for citing articles in:
ISI Web of Science (1)
Google Scholar
Articles by Caballero-Alomar, C.
Articles by Puig-Parellada, P.
Articles citing this Article
PubMed
PubMed Citation
Articles by Caballero-Alomar, C.
Articles by Puig-Parellada, P.


HOME HELP FEEDBACK SUBSCRIPTIONS ARCHIVE SEARCH TABLE OF CONTENTS
Visit Other APS Journals Online
Copyright © 2003 by the American Physiological Society.