Biomedical subjects

K M Sanders

Publications and source records attributed to K M Sanders.

At least 91 records · Page 5Linked to original sources

PubMed1999

Development and plasticity of interstitial cells of Cajal.

Interstitial cells of Cajal (ICC) are the pacemakers in gastrointestinal (GI) muscles, and these cells also mediate or transduce inputs from the enteric nervous system. Different classes of ICC are involved in pacemaking and neurotransmission. ICC express specific ionic conductances that make them unique in their ability to generate and propagate slow waves in GI muscles or transduce neural inputs. Much of what we know about the function of ICC comes from developmental studies that were made possible by the discoveries that ICC express c-kit and proper development of ICC depends upon signalling via the Kit receptor pathway. Manipulating Kit signalling with reagents to block the receptor or downstream signalling pathways or by using mutant mice in which Kit or its ligand, stem cell factor, are defective has allowed novel studies into the specific functions of the different classes of ICC in several regions of the GI tract. Kit is also a surface antigen that can be used to conveniently label ICC in GI muscles. Immunohistochemical studies using Kit antibodies have expanded our knowledge about the ICC phenotype, the structure of ICC networks, the interactions of ICC with other cells in the gut wall, and the loss of ICC in some clinical disorders. Preparations made devoid of ICC have also allowed analysis of the consequences of losing specific classes of ICC on GI motility. This review describes recent advances in our knowledge about the development and plasticity of ICC and how developmental studies have contributed to our understanding of the functions of ICC. We have reviewed the clinical literature and discussed how loss or defects in ICC affect GI motor function.

Animals↗

PubMed1999

Cellular and molecular basis for electrical rhythmicity in gastrointestinal muscles.

Regulation of gastrointestinal (GI) motility is intimately coordinated with the modulation of ionic conductance expressed in GI smooth muscle and nonmuscle cells. Interstitial cells of Cajal (ICC) act as pacemaker cells and possess unique ionic conductances that trigger slow wave activity in these cells. The slow wave mechanism is an exclusive feature of ICC: Smooth muscle cells may lack the basic ionic mechanisms necessary to generate or regenerate slow waves. The molecular identification of the components for these conductances provides the foundation for a complete understanding of the ionic basis for GI motility. In addition, this information will provide a basis for the identification or development of therapeutics that might act on these channels. It is much easier to study these conductances and develop blocking drugs in expression systems than in native GI muscle cells. This review focuses on the relationship between ionic currents in native GI smooth muscle cells and ICC and their molecular counterparts.

Animals↗

PubMed1999

Review article: effects of the 5-HT3 receptor antagonist alosetron on neuromuscular transmission in canine and human intestinal muscle.

BACKGROUND: Currently, therapeutic treatments for irritable bowel syndrome fail to produce significant clinical results. We hypothesized that alosetron, a selective 5-HT3 antagonist, may provide symptomatic relief in irritable bowel syndrome patients through a decrease in the amplitude of gastrointestinal contractions. AIM: To determine the in vitro effect of alosetron on neuromuscular transmission in the canine and human jejunal and colonic muscularis externa. RESULTS: Alosetron diminished electrical field-stimulated (EFS) contractions recorded from muscles of the canine and human small and large intestines. Mechanistically, the diminished EFS response could be explained by the ability of alosetron to decrease the fractional release of 14C-choline radiolabelled acetylcholine evoked by EFS from human jejunal muscle. The inhibition of EFS contractions was not limited to atropine-sensitive events, as non-cholinergic excitatory EFS evoked contractions were also inhibited. Additionally, alosetron at high concentrations (> 30 microM) directly altered bethanechol stimulated contractions. CONCLUSION: Caution must be used in the interpretation of these data because significant alterations in EFS-induced contractions were only observed with large pharmacological concentrations of alosetron, and the response was not selective for cholinergically-mediated excitatory neuromuscular transmission.

Animals↗

PubMed1998

Spontaneous electrical rhythmicity in cultured interstitial cells of cajal from the murine small intestine.

1. Interstitial cells of Cajal (ICC) are pacemaker cells in the small bowel, and therefore this cell type must express the mechanism responsible for slow wave activity. Isolated ICC were cultured for 1-3 days from the murine small intestine and identified with c-Kit-like immunoreactivity (c-Kit-LI). 2. Electrical recordings were obtained from cultured ICC with the whole-cell patch clamp technique. ICC were rhythmically active, producing regular slow wave depolarizations with waveforms and properties similar to slow waves in intact tissues. 3. Spontaneous activity of c-Kit-LI cells was inhibited by reduced extracellular Na+, gadolinium, and reduced extracellular Ca2+. The activity was not affected by nisoldipine. Voltage clamp studies showed rhythmic inward currents that were probably responsible for the slow wave activity. The current-voltage relationship showed that the spontaneous currents reversed at about +17 mV. These observations are consistent with the involvement of a non-selective cation current in the generation of slow waves, but do not rule out contributions from other conductances or transporters. 4. A Ba2+-sensitive inwardly rectifying K+ current in c-Kit-LI cells that may be involved in slow wave repolarization and maintenance of a negative potential between slow waves was also found. Similar pharmacology was observed in studies of intact murine intestinal muscles. 5. Cultured ICC may be a useful model for studying the properties and pharmacology of some of the ionic conductances involved in spontaneous rhythmicity in the gastrointestinal tract.

Animals↗

PubMed1998

Control of motility patterns in the human colonic circular muscle layer by pacemaker activity.

1. This study characterized the electrical and mechanical activities of human colonic muscle strips obtained from either the ascending, descending or sigmoid colon of patient volunteers during elective colon resections. 2. Rhythmic contractile activity was observed in colonic circular muscle strips in the absence of external stimuli. This activity persisted in the presence of atropine, phentolamine, propranolol, tetrodotoxin and Nomega-nitro-L-arginine but was abolished by nifedipine. 3. The activity of whole circular muscle (WCM) was compared with that of the myenteric half (MCM), the submucosal half (SCM) and the interior (ICM) of the circular muscle layer. WCM exhibited a prominent 2-4 contractions min-1 contractile pattern which was also present in strips of SCM. In contrast, MCM and ICM exhibited slow (0.3-0.6 contractions min-1), long duration contractions with superimposed higher frequency contractions (17-18 contractions min-1). 4. Resting membrane potential (Vm), recorded at various positions through the thickness of WCM strips did not differ and averaged -50 mV. 5. Slow waves were observed in 83 % of muscles. They averaged 12 mV in amplitude, 9.4 s in duration and had a frequency of 2-4 contractions min-1. Slow waves were greatest in amplitude near the submucosal edge and decreased with distance away from this edge. Each slow wave was associated with a transient contraction. 6. Near the myenteric edge, rapid fluctuations of Vm with a mean frequency of 18 contractions min-1 were recorded in 67 % of muscles. Spiking activity was common and was superimposed upon slow waves and rapid Vm fluctuations. 7. In summary, slow waves were identified in the human colonic circular muscle layer which arise at or near the submucosal edge. These electrical events give rise to a 2-4 contractions min-1 contractile rhythm which is characteristic of the intact muscle layer. Thus, the nature and spatial organization of pacemaker activity in the human colon bears significant resemblance to other animal models, such as the dog and pig.

Adolescent↗

PubMed1998

Dissociation between electrical and mechanical responses to nitrergic stimulation in the canine gastric fundus.

1. We examined the relationships between membrane potential, intracellular [Ca2+] ([Ca2+]i), and tension in muscles of the canine gastric fundus in response to nitrergic stimulation by NO donors and electrical field stimulation (EFS) of intrinsic enteric inhibitory neurons when adrenergic and cholinergic responses were blocked. 2. NO donors reduced [Ca2+]i and tension in a concentration-dependent manner. A close relationship was noted between these parameters. 3. In terms of the [Ca2+] vs. force relationship, relaxation responses to EFS differed from responses to NO donors. EFS resulted in smaller decreases in [Ca2+]i to produce a given relaxation compared with responses to NO donors. Thus, muscles stimulated with EFS were less sensitive to [Ca2+]i than muscles stimulated with exogenous NO. 4. When membrane potential, [Ca2+]i and tension were monitored simultaneously in the same muscles, a temporal dissociation was noted between the electrical responses and changes in [Ca2+]i and tension. Brief electrical responses were associated with more sustained changes in [Ca2+]i and tension. 5. Further dissociation between electrical and mechanical effects was noted. Changes in [Ca2+]i and tension caused by sodium nitroprusside and EFS were blocked by arginine analogues and by oxyhaemoglobin, but electrical responses were unaffected. 1H-[1,2,4]oxadiazolo[4, 3-a]quinoxalin-1-one (ODQ), an inhibitor of soluble guanylyl cyclase, also blocked the effects of nitrergic stimulation on [Ca2+]i and tension, without affecting hyperpolarization. Thus, in the presence of continued hyperpolarization, the reductions in [Ca2+]i and tension caused by nitrergic stimulation were blocked. 6. Block of hyperpolarization in response to nitrergic stimulation with tetrapentylammonium chloride (TPEA) had relatively little effect on the [Ca2+]i and tension responses. Thus, hyperpolarization is not required for nitrergic effects on [Ca2+]i and tension. 7. In summary, reduction in [Ca2+]i and tension in response to nitrergic stimulation of the canine gastric fundus does not depend upon electrical hyperpolarization. Non-electrical mechanisms such as enhanced uptake of Ca2+ by the sarcoplasmic reticulum or reduction in the Ca2+ sensitivity of the contractile apparatus may be the primary mechanisms mediating nitrergic responses in these muscles.

Animals↗

PubMed1998

Basal activation of ATP-sensitive potassium channels in murine colonic smooth muscle cell.

The function and molecular expression of ATP-sensitive potassium (KATP) channels in murine colonic smooth muscle was investigated by intracellular electrical recording from intact muscles, patch-clamp techniques on isolated smooth muscle myocytes, and reverse transcription polymerase chain reaction (RT-PCR) on isolated cells. Lemakalim (1 microM) caused hyperpolarization of intact muscles (17. 2 +/- 3 mV). The hyperpolarization was blocked by glibenclamide (1-10 microM). Addition of glibenclamide (10 microM) alone resulted in membrane depolarization (9.3 +/- 1.7 mV). Lemakalim induced an outward current of 15 +/- 3 pA in isolated myocytes bathed in 5 mM external K+ solution. Application of lemakalim to cells in symmetrical K+ solutions (140/140 mM) resulted in a 97 +/- 5 pA inward current. Both currents were blocked by glibenclamide (1 microM). Pinacidil (1 microM) also activated an inwardly rectifying current that was insensitive to 4-aminopyridine and barium. In single-channel studies, lemakalim (1 microM) and diazoxide (300 microM) increased the open probability of a 27-pS K+ channel. Openings of these channels decreased with time after patch excision. Application of ADP (1 mM) or ATP (0.1 mM) to the inner surface of the patches reactivated channel openings. The conductance and characteristics of the channels activated by lemakalim were consistent with the properties of KATP. RT-PCR demonstrated the presence of Kir 6.2 and SUR2B transcripts in colonic smooth muscle cells; transcripts for Kir 6.1, SUR1, and SUR2A were not detected. These molecular studies are the first to identify the molecular components of KATP in colonic smooth muscle cells. Together with the electrophysiological experiments, we conclude that KATP channels are expressed in murine colonic smooth muscle cells and suggest that these channels may be involved in dual regulation of resting membrane potential, excitability, and contractility.

Adenosine Diphosphate↗

PubMed1998

Interstitial cells of Cajal mediate enteric inhibitory neurotransmission in the lower esophageal and pyloric sphincters.

BACKGROUND & AIMS: Previous studies have suggested that a specific class of interstitial cells of Cajal (ICC) act as mediators in nitrergic inhibitory neurotransmission. The aim of this investigation was to examine the role of intramuscular ICC (IC-IM) in neurotransmission in the murine lower esophageal (LES) and pyloric sphincters (PS). METHODS: Immunohistochemistry and electrophysiology were used to study the distribution and role of IC-IM. RESULTS: The LES and PS contain spindle-shaped IC-IM, which form close relationships with nitric oxide synthase-containing nerve fibers. The PS contains ICC within the myenteric plexus and c-Kit immunopositive cells along the submucosal surface of the circular muscle. IC-IM were absent in the LES and PS of c-kit (W/Wv) mutant mice. Using these mutants, we tested whether IC-IM mediate neural inputs in the LES and PS. Although the distribution of inhibitory nerves was normal in W/Wv animals, NO-dependent inhibitory neurotransmission was reduced. Hyperpolarizations to sodium nitroprusside were also attenuated in W/Wv animals. CONCLUSIONS: The data suggest that IC-IM play an important role in NO-dependent neurotransmission in the LES and PS. IC-IM may be the effectors that transduce NO signals into hyperpolarizing responses. Loss of IC-IM may interfere with relaxations and normal motility in these sphincters.

Animals↗

PubMed1998

Identification of neurons that express stem cell factor in the mouse small intestine.

BACKGROUND & AIMS: Enteric neurons in the murine intestine express stem cell factor (SCF), which may provide an important signal in the development of the interstitial cells of Cajal (ICC). Our aim was to identify the subpopulation(s) of myenteric neurons that express SCF. METHODS: Myenteric plexus preparations from postnatal SCF-lacZ mice were processed for beta-galactosidase histochemistry followed by immunohistochemistry. RESULTS: Approximately 60% of the nitric oxide synthase-immunoreactive neurons, which projected to myenteric ganglia and to circular muscle, expressed SCF, and more than 80% of the calbindin-immunoreactive neurons, which projected exclusively to myenteric ganglia, expressed SCF. A small subpopulation of calretinin-immunoreactive neurons expressed SCF transiently. Many of the remainder of SCF-expressing neurons were choline acetyltransferase immunoreactive, but their projections are unknown. CONCLUSIONS: SCF-expressing neurons that project within the myenteric plexus may be an important source of SCF for the development of Kit-expressing ICC at this level. The only possible neuronal source of SCF for the ICC of the deep muscular plexus is a subpopulation of nitric oxide synthase-immunoreactive neurons.

Animals↗

PubMed1998

Is malnutrition overdiagnosed in older hospitalized patients? Association between the soluble interleukin-2 receptor and serum markers of malnutrition.

BACKGROUND: Many researchers have speculated that markers of malnutrition such as albumin, prealbumin, cholesterol, and transferrin are influenced by inflammation. The mechanism of this interaction has not been well understood. METHODS: This was a prospective cross-sectional study. We evaluated 72 male patients older than 60 years admitted to a geriatric rehabilitation unit. Subjects with severe hepatic or renal diseases were excluded. We measured body mass index, caloric intake, serum albumin, prealbumin, cholesterol, transferrin, hemoglobin, and total lymphocyte count. To detect inflammation, we measured C-reactive protein, Westergren sedimentation rate, fibrinogen, and cytokines including tumor necrosis factor-alpha (TNF-alpha), interleukin-1 beta (IL-1 beta), IL-6, IL-2, and the soluble IL-2 receptor. RESULTS: Soluble IL-2 receptor was negatively associated with albumin (r = -.479, p < .0001), prealbumin (r = -.520, p = < .0001), cholesterol (r = -.487, p = .0001), transferrin (r = -.455, p = .0002), and hemoglobin (r = -.371, p = .002). TNF-alpha, IL-1 beta, IL-6, and IL-2 were not associated with these measures. CONCLUSIONS: Inflammation increases the incidence of hypoalbuminemia and hypocholesterolemia, potentially leading to overdiagnosis of malnutrition. We suggest that albumin, cholesterol, prealbumin, and transferrin be used with caution when assessing the nutritional status of older hospitalized patients. In the future, soluble IL-2 receptor levels might be used to correct for the impact of inflammation on these markers of malnutrition.

Aged↗

PubMed1998

Functional and molecular identification of a novel chloride conductance in canine colonic smooth muscle.

Swelling-activated or volume-sensitive Cl- currents are found in numerous cell types and play a variety of roles in their function; however, molecular characterization of the channels is generally lacking. Recently, the molecular entity responsible for swelling-activated Cl- current in cardiac myocytes has been identified as ClC-3. The goal of our study was to determine whether such a channel exists in smooth muscle cells of the canine colon using both molecular biological and electrophysiological techniques and, if present, to characterize its functional and molecular properties. We hypothesized that ClC-3 is present in colonic smooth muscle and is regulated in a manner similar to the molecular entity cloned from heart. Indeed, the ClC-3 gene was expressed in colonic myocytes, as demonstrated by reverse transcriptase polymerase chain reaction performed on isolated cells. The current activated by decreasing extracellular osmolarity from 300 to 250 mosM was outwardly rectifying and dependent on the Cl- gradient. Current magnitude increased and reversed at more negative potentials when Cl- was replaced by I- or Br-. Tamoxifen ([Z]-1-[p-dimethylaminoethoxy-phenyl]-1,2-diphenyl-1-butene; 10 microM) and DIDS (100 microM) inhibited the current, whereas 25 microM niflumic acid, 10 microM nicardipine, and Ca2+ removal had no effect. Current was inhibited by 1 mM extracellular ATP in a voltage-dependent manner. Cl- current was also regulated by protein kinase C, as phorbol 12,13-dibutyrate (300 nM) decreased Cl- current magnitude, while chelerythrine chloride (30 microM) activated it under isotonic conditions. Our findings indicate that a current activated by hypotonic solution is present in colonic myocytes and is likely mediated by ClC-3. Furthermore, we suggest that the ClC-3 may be an important mechanism controlling depolarization and contraction of colonic smooth muscle under conditions that impose physical stress on the cells.

4,4'-Diisothiocyanostilbene-2,2'-Disulfonic Acid↗

PubMed1998

G protein-coupled receptors in gastrointestinal physiology. IV. Neural regulation of gastrointestinal smooth muscle.

G protein-coupled receptors receive many of the neural, hormonal, and paracrine inputs to gastrointestinal (GI) smooth muscle cells. This article examines the major G protein-coupled receptors, G proteins, and effectors that mediate responses to enteric neuromuscular transmitters. Excitatory transmitters primarily couple through Gq/11 and Gi/Go proteins and elicit responses via formation of inositol trisphosphate and diacylglycerol and inhibition of adenylyl cyclase. Several inhibitory transmitters couple through Gs and activation of adenylyl cyclase. There are interesting examples, however, of inhibitory transmitters apparently using pathways regulated by Gq/11 to elicit responses by localized Ca2+ release and activation of Ca2+-dependent ion channels. G protein-coupled receptors may also be differentially expressed by smooth muscle cells and interstitial cells of Cajal, which may increase the diversity of responses and allow specialized innervation of GI muscle tissues.

Adenylyl Cyclases↗

PubMed1998

The exclusion of high trauma fractures may underestimate the prevalence of bone fragility fractures in the community: the Geelong Osteoporosis Study.

Fractures associated with severe trauma are generally excluded from estimates of the prevalence of osteoporotic fractures in the community. Because the degree of trauma is difficult to quantitate, low bone mass may contribute to fractures following severe trauma. We ascertained all fractures in a defined population and compared the bone mineral density (BMD) of women who sustained fractures in either "low" or "high" trauma events with the BMD of a random sample of women from the same population. BMD was measured by dual-energy X-ray absorptiometry and expressed as a standardized deviation (Z score) adjusted for age. The BMD Z scores (mean +/- SEM) were reduced in both the low and high trauma groups, respectively: spine-posterior-anterior (-0.50 +/- 0.05 and -0.21 +/- 0.08), spine-lateral (-0.28 +/- 0.06 and -0.19 +/- 0.10), femoral neck (-0.42 +/- 0.04 and -0.26 +/- 0.09), Ward's triangle (-0.44 +/- 0.04 and -0.28 +/- 0.08), trochanter (-0.44 +/- 0.05 and -0.32 +/- 0.08), total body (-0.46 +/- 0.06 and -0.32 +/- 0.08), ultradistal radius (-0.47 +/- 0.05 and -0.42 +/- 0.07), and midradius (-0.52 +/- 0.06 and -0.33 +/- 0.09). Except at the PA spine, the deficits were no smaller in the high trauma group. Compared with the population, the age-adjusted odds ratio for osteoporosis (t-score < -2.5) at one or more scanning sites was 3.1 (95% confidence interval 1.9, 5.0) in the high trauma group and 2.7 (1.9, 3.8) in the low trauma group. The data suggest that the exclusion of high trauma fractures in women over 50 years of age may result in underestimation of the contribution of osteoporosis to fractures in the community. Bone density measurement of women over 50 years of age who sustain fractures may be warranted irrespective of the classification of trauma.

Absorptiometry, Photon↗

PubMed1997

Development of electrical rhythmicity in the murine gastrointestinal tract is specifically encoded in the tunica muscularis.

1. Interstitial cells of Cajal (ICCs) have been identified as pacemaker cells in the gastrointestinal (GI) tracts of vertebrates. We have studied the development of ICCs in pacemaker regions and the onset of electrical rhythmicity in the gastric antrum, small bowel and proximal colon of the mouse. 2. ICCs, as detected by c-Kit immunofluorescence, were found during embryogenesis in regions of the GI tract that eventually become pacemaker areas. Prior to birth, these cells were organized into well-structured networks, and by the end of the embryonic period the morphology of ICC networks in pacemaker regions appeared very similar to that observed in adult animals. 3. Electrical rhythmicity was recorded prior to birth (by E18) in the proximal GI tract (stomach and jejunum), and this activity developed to adult-like behaviour within a week after birth. In the ileum and proximal colon rhythmicity developed after birth, and adult-like characteristics were apparent within the first week. 4. Post-junctional responses of smooth muscles to neural inputs could be recorded at birth, and stimulation of intrinsic nerves often led to oscillatory activity resembling slow waves for up to several minutes following brief stimuli. Nerve stimulation augmented spontaneous activity in the proximal portions of the GI tract and elicited rhythmic activity temporarily in quiescent tissues of the distal GI tract. 5. ICCs and rhythmicity developed in an apparently normal manner in tissues isolated at birth and placed in organ culture. These data suggest that the tunica muscularis provides a suitable microenvironment for the development of ICCs and rhythmicity without the need for extrinsic stimuli. 6. Treatment of small intestinal tissues taken from embryos at E15 with neutralizing c-Kit antibodies abolished ICC development and the organization of ICCs into networks that typically occurs during the late embryonic period. Treatment of muscles taken from newborn animals with c-Kit antibodies blocked postnatal development of ICCs, disrupted already established and functional ICC networks, and rendered muscles electrically quiescent. 7. In summary, ICC networks develop in the pacemaker regions of the murine GI tract before birth. Development and organization of ICCs of the myenteric plexus region into networks precedes the development of electrical rhythmicity. Post-natal development of electrical rhythmicity is mainly characterized by enhancement of the amplitude and frequency of slow waves. The development of ICCs and electrical rhythmicity persists in vitro. ICCs appear to be necessary for the initiation of electrical rhythmicity. These findings provide further evidence for the pacemaker role of ICCs.

Animals↗

PubMed1997

Basal release of nitric oxide induces an oscillatory motor pattern in canine colon.

1. The consequences of intrinsic, basal nitric oxide release on electrical and contractile activity of canine proximal colon were examined. Membrane potential and contraction were simultaneously recorded from the circular muscle in the presence of drugs to block adrenergic and cholinergic responses. 2. Electrical slow waves were recorded from muscle cells near the submucosal surface of the circular layer. Spontaneous contractions were initiated by each slow wave. Contractile amplitude increased 1.9-fold when nerves were blocked with tetrodotoxin (TTX, 1 microM). 3. Muscle cells near the myenteric surface displayed myenteric potential oscillations (MPOs) averaging 16 cycles per minute (c.p.m.) in frequency and 10 mV in amplitude. Twenty-five per cent of muscles displayed an additional slow, neurogenic oscillation (mean frequency, 1 c.p.m.; amplitude, 14 mV) superimposed upon the MPO rhythm. 4. The nitric oxide (NO) synthase inhibitor N omega -nitro-L-arginine (L-NA, 100 microM; n = 16) abolished neurogenic oscillations, depolarized cells, and increased MPO upstroke velocity, amplitude and frequency. The actions of L-NA were mimicked by N omega-nitro-L-arginine methylester (L-NAME, 100 microM) and oxyhaemoglobin (3%). 5. Spontaneous contractions were increased 2.3-fold by L-NA, and TTX had no effect on contractions after addition of L-NA. 6. The NO-donor sodium nitroprusside (SNP, 1 microM) reversed the electrical and mechanical effects of L-NA and initiated slow oscillations similar to the neurogenic oscillations. Slow oscillations were also evoked with S-nitroso-N-acetylpenicillamine (SNAP, 1 microM). The effects of NO donors were blocked by oxyhaemoglobin. 7. Slow electrical oscillations could not be elicited by SNP after removal of a thin strip of circular muscle along the myenteric edge. 8. These data suggest that the spontaneous electrical and contractile activity of the proximal colon is tonically suppressed by basal release of NO. Basal NO causes an oscillatory pattern of electrical and mechanical activity. This activity does not require patterned firing of nerves; rather a continuous, low level release of NO would be capable of producing the neurogenic oscillatory behaviour. The slow oscillatory activity depends upon the presence of the myenteric region of the circular muscle layer, which contains cell bodies of enteric neurons and interstitial cells of Cajal.

Animals↗

PubMed1997

Interstitial cells of Cajal in the guinea-pig gastrointestinal tract as revealed by c-Kit immunohistochemistry.

Interstitial cells of Cajal (ICC) of various morphologies have been described in the gastrointestinal (GI) tracts of mammals. Different classes of ICC are likely to have different functional roles. ICC of the mouse GI tract have been shown to express c-kit, a proto-oncogene that codes for a receptor tyrosine kinase. We have studied the distribution of ICC within the guinea pig GI tract using antibodies to c-Kit protein and immunohistochemical techniques. c-Kit-like immunoreactivity revealed at least 6 types of ICC: (1) intramuscular ICC (IC-IM1) that lie within the muscle layers of the esophagus, stomach, and cecum, (2) ICC within the myenteric plexus region (IC-MY1) in the corpus, antrum, small intestine, and colon, (3) ICC that populate the deep muscular plexus of the small intestine (IC-DMP), (4) ICC at the submucosal surface of the circular muscle layer in the colon (IC-SM), (5) stellate ICC that are closely associated with the myenteric plexus (IC-MY2) and orientated toward the longitudinal muscle layer in the colon, and (6) branching intramuscular ICC (IC-IM2) in the proximal colon within the circular and longitudinal muscle layers. c-Kit immunohistochemistry appears to be an excellent and selective technique for labeling ICC of the guinea-pig GI tract. Labeling of these cells at the light-microscopic level provides an opportunity for characterizing the distribution, density, organization, and relationship between ICC and other cell types.

Animals↗

PubMed1997

Development of c-Kit-positive cells and the onset of electrical rhythmicity in murine small intestine.

BACKGROUND & AIMS: Little is known about the development of interstitial cells (ICs), yet these cells are important in electrical rhythmicity and neurotransmission in the gastrointestinal tract. This study characterized the development of ICs and the onset of electrical rhythmicity in the murine intestine. METHODS: Antibodies against c-Kit (e.g., the receptor for stem cell factor) were used to label ICs of the small intestines of embryos and neonatal mice. Labels for enteric neuroblasts and smooth muscle cells were used to study neighboring cells. Development was examined also with electron microscopy and electrophysiological techniques. RESULTS: c-Kit-like immunoreactivity (c-Kit-LI) was detected in gastrointestinal tissues at embryonic day 12.5. Labeled cells were distributed along the outer perimeter of the intestine and had morphological features of neither smooth muscle cells nor ICs. Cells with c-Kit-LI were nonneural and seemed to be common precursors for longitudinal muscle cells and ICs of the myenteric plexus region (IC-MY). Longitudinal muscle cells lost c-Kit by E18, whereas IC-MY continued c-Kit expression into adulthood. Electrical rhythmicity developed after IC-MY, and longitudinal muscle cells became separate entities. ICs in the deep muscular plexus region developed after birth. CONCLUSIONS: ICs have a nonneural origin. Common precursors yield IC-MY and longitudinal muscle cells. Development of IC-MY correlates with the initiation of electrical rhythmicity.

Animals↗

PubMed1997

Regulation of citrulline recycling in nitric oxide-dependent neurotransmission in the murine proximal colon.

1. We investigated the contribution of nitric oxide (NO) to inhibitory neuromuscular transmission in murine proximal colon and the possibility that citrulline is recycled to arginine to maintain the supply of substrate for NO synthesis. 2. Intracellular microelectrode recordings were made from circular smooth muscle cells in the presence of nifedipine and atropine (both 1 microM). Electrical field stimulation (EFS, 0.3-20 Hz) produced inhibitory junction potentials (i.j.ps) composed of an initial transient hyperpolarization (fast component) followed by a slow recovery to resting potential (slow component). 3. L-Nitro-arginine-methyl ester (L-NAME, 100 microM) selectively abolished the slow component of i.j.ps. The effects of L-NAME were reversed by L-arginine (0.2-2 mM) but not by D-arginine (2 mM). Sodium nitroprusside (an NO donor, 1 microM) reversibly hyperpolarized muscle cells. This suggests that NO mediates the slow component of i.j.ps. 4. L-Citrulline (0.2 mM) also reversed the effects of L-NAME, and this action was maintained during sustained exposures to L-citrulline (0.2 mM). This may reflect intraneuronal recycling of L-citrulline to L-arginine. 5. Higher concentrations of L-citrulline (e.g. 2 mM) had time-dependent effects. Brief exposure (15 min) reversed the effects of L-NAME, but during longer exposures (30 min) the effects of L-NAME gradually returned. In the continued presence of L-citrulline, L-arginine (2 mM) readily restored nitrergic transmission, suggesting that during long exposures to high concentrations of L-citrulline, the ability to generate arginine from citrulline was reduced. 6. Aspartate (2 mM) had no effect on i.j.ps, the effects of L-NAME, or the actions of L-citrulline in the presence of L-NAME, L-Citrulline (0.2-2 mM) alone had no effect on i.j.ps under control conditions. 7. S-methyl-L-thiocitrulline (10 microM), a novel NOS inhibitor, blocked the slow component of i.j.ps. The effects of this inhibitor were reversed by L-arginine (2 mM), but not by L-citrulline (2 mM). 8. These results suggest that i.j.ps in the murine colon result from release of multiple inhibitory neurotransmitters. NO mediates a slow component of enteric inhibitory neurotransmission. Recycling of L-citrulline to L-arginine may sustain substrate concentrations in support of NO synthesis and this pathway may be inhibited when concentrations of L-citrulline are elevated.

Animals↗