scholarly journals Adrenergic activation of electrogenic K+ secretion in guinea pig distal colonic epithelium: desensitization via the Y2-neuropeptide receptor

2009 ◽  
Vol 297 (2) ◽  
pp. G278-G291 ◽  
Author(s):  
Jin Zhang ◽  
Susan T. Halm ◽  
Dan R. Halm
Keyword(s):  
Guinea Pig ◽  
Peptide Yy ◽  
Short Circuit ◽  
In Vitro Release ◽  
Surface Epithelium ◽  
Prostaglandin E ◽  
Bathing Solution ◽  
Ussing Chambers ◽  
Adrenergic Activation

Adrenergic activation of electrogenic K+ secretion in isolated mucosa from guinea pig distal colon was desensitized by peptide-YY (PYY). Addition of PYY or neuropeptide-Y (NPY) to the bathing solution of mucosae in Ussing chambers suppressed the short-circuit current ( Isc) corresponding to electrogenic Cl− secretion, whether stimulated by epinephrine (epi), prostaglandin-E2 (PGE2), or carbachol (CCh). Neither peptide markedly inhibited the large transient component of synergistic secretion (PGE2 + CCh). Sustained Cl− secretory Isc was inhibited ∼65% by PYY or NPY, with IC50s of 4.1 ± 0.9 nM and 9.4 ± 3.8 nM, respectively. This inhibition was eliminated by BIIE0246, an antagonist of the Y2-neuropeptide receptor (Y2-NpR), but not by Y1-NpR antagonist BVD10. Adrenergic sensitivity for activation of K+ secretion in the presence of Y2-NpR blockade by BIIE0246 was (EC50s) 2.9 ± 1.2 nM for epi and 13.3 ± 1.0 nM for norepinephrine, approximately fourfold greater than in the presence of PYY. Expression of mRNA for both Y1-NpR and Y2-NpR was indicated by RT-PCR of RNA from colonic mucosa, and protein expression was indicated by immunoblot. Immunoreactivity (ir) for Y1-NpR and Y2-NpR was distinct in basolateral membranes of columnar epithelial cells in the crypts of Lieberkühn as well as intercrypt surface epithelium. Adrenergic nerves in proximity with crypts were detected by ir for dopamine-β-hydroxylase, and a portion of these nerves also contained NPYir. BIIE0246 addition increased secretagog-activated Isc, consistent with in vitro release of either PYY or NPY. Thus PYY and NPY were able to suppress Cl− secretory capacity and desensitize the adrenergic K+ secretory response, providing a direct inhibitory counterbalance against secretory activation.

scholarly journals Adrenergic activation of electrogenic K+ secretion in guinea pig distal colonic epithelium: involvement of β1- and β2-adrenergic receptors

2009 ◽  
Vol 297 (2) ◽  
pp. G269-G277 ◽  
Author(s):  
Jin Zhang ◽  
Susan T. Halm ◽  
Dan R. Halm
Keyword(s):  
Epithelial Cells ◽  
Guinea Pig ◽  
Short Circuit ◽  
Surface Epithelium ◽  
Receptor Subtype ◽  
Receptor Subtypes ◽  
Bathing Solution ◽  
Adrenergic Stimulation ◽  
Ussing Chambers ◽  
Adrenergic Activation

Adrenergic stimulation of electrogenic K+ secretion in isolated mucosa from guinea pig distal colon required activation of two β-adrenergic receptor subtypes (β-AdrR). Addition of epinephrine (epi) or norepinephrine (norepi) to the bathing solution of mucosae in Ussing chambers increased short-circuit current ( Isc) and transepithelial conductance ( Gt), consistent with this cation secretion. A β-adrenergic classification was supported by propranolol antagonism of this secretory response and the lack of effect by the α-AdrR antagonists BE2254 (α1-AdrR) and yohimbine (α2-AdrR). Subtype-selective antagonists CGP20712A (β1-AdrR), ICI-118551 (β2-AdrR), and SR59320A (β3-AdrR) were relatively ineffective at inhibiting the epi-stimulated Isc response. In combination, CGP20712A and ICI-118551 inhibited the response, which supported a synergistic action by β1-AdrR and β2-AdrR. Expression of mRNA for both β1-AdrR and β2-AdrR was indicated by RT-PCR of RNA from colonic epithelial cells. Protein expression was indicated by immunoblot showing bands at molecular weights consistent with monomers and oligomers. Immunoreactivity (ir) for β1-AdrR and β2-AdrR was prominent in basolateral membranes of columnar epithelial cells in the crypts of Lieberkühn as well as intercrypt surface epithelium. Cells in the pericryptal sheath also had β1-AdrRir but did not have discernable β2-AdrRir. The adrenergic sensitivity of K+ secretion measured by Isc and Gt was relatively low as indicated by EC50s of 41 ± 7 nM for epi and 50 ± 14 nM for norepi. Adrenergic activation of electrogenic K+ secretion required the involvement of both β1-AdrR and β2-AdrR, occurring with an agonist sensitivity reduced compared with reported values for either receptor subtype.

Bicarbonate secretion by rabbit proximal colon

1986 ◽  
Vol 251 (4) ◽  
pp. G436-G445 ◽  
Author(s):  
S. K. Sullivan ◽  
P. L. Smith
Keyword(s):  
Transport Process ◽  
Short Circuit ◽  
Proximal Colon ◽  
Bathing Solution ◽  
Bicarbonate Secretion ◽  
Rabbit Ileum ◽  
Ussing Chambers ◽  
Ph Stat ◽  
Dependent Mechanism

Stripped segments of proximal colon (1-6 cm distal to the ampulla caecalis coli) were studied in vitro in Ussing chambers under short-circuit conditions using the pH-stat technique. With glucose and HCO3-CO2 present in the serosal bathing solution only, proximal colon alkalinizes the luminal bathing solution at a rate of 2.1 +/- 0.2 mu eq X h-1 X cm-2 (n = 36). With HCO3-CO2 present in the luminal bathing solution alone, proximal colon does not significantly acidify or alkalinize the serosal bathing solution. Addition of glucose (10 mM) to the luminal bathing solution abolished luminal alkalinization. Removal of HCO3 and CO2 from the serosal bathing solution or replacement of O2 with N2 also abolished luminal alkalinization. Acetazolamide (0.1 mM) added to both bathing solutions did not alter the rate of luminal alkalinization. Ion-replacement studies revealed that the alkalinization process was highly dependent on the presence of Na in the bathing solutions and much less dependent on the presence of Cl. Furthermore, ouabain (0.1 mM) significantly reduced luminal alkalinization. As in rabbit ileum, serosal epinephrine (0.1 mM) did not alter luminal alkalinization but increased serosal alkalinization by a Na-dependent mechanism. These results suggest that luminal alkalinization results from a Na-dependent, active transcellular HCO3 transport process and that a Na-dependent HCO3 absorptive process is activated by adrenergic stimuli.

Effect of sulfidopeptide leukotrienes D4 and E4 on ileal ion transport in vitro in the rat and rabbit

1988 ◽  
Vol 255 (2) ◽  
pp. G175-G183 ◽  
Author(s):  
P. L. Smith ◽  
D. P. Montzka ◽  
G. P. McCafferty ◽  
M. A. Wasserman ◽  
J. D. Fondacaro
Keyword(s):  
Short Circuit ◽  
Short Circuit Current ◽  
Bathing Solution ◽  
Na Transport ◽  
Rabbit Ileum ◽  
Rat Ileum ◽  
Ussing Chambers ◽  
Meclofenamic Acid ◽  
Acid Metabolites

Effects of leukotrienes D4 and E4 (LTD4 and LTE4) on electrolyte transport were examined, employing stripped segments of rat and rabbit ileum mounted in Ussing chambers. Addition of LTD4 or LTE4 to the serosal but not the mucosal bathing solution elicited a transient increase in short-circuit current (Isc) with maximal responses seen at 10(-5) M and 10(-8) M in rat and rabbit respectively and a sustained decrease in transepithelial conductance (Gt) in the rat only. In the rat, Cl replacement, reduction of bathing solution [Ca2+] to 1 microM or pretreatment with 1 microM indomethacin or meclofenamic acid inhibited the LTD4- or LTE4-induced Isc changes with no effect on the decrease in Gt. LTD4 (10 microM) transiently increased net Cl secretion and produced a sustained decrease in both unidirectional and net Na transport and mucosal-to-serosal Cl flux in rat ileum. The decrease in unidirectional Na fluxes is accounted for predominantly by a change in the potential independent flux of Na. These results suggest that the increase in Isc in both rat and rabbit is mediated by arachidonic acid metabolites, whereas the decrease in Gt and net Na absorption in rat ileum is mediated by a cyclooxygenase-independent pathway.

Structural and functional changes by ethanol on in vitro guinea pig gastric mucosa

1986 ◽  
Vol 251 (4) ◽  
pp. G518-G528 ◽  
Author(s):  
M. J. Rutten ◽  
S. Ito
Keyword(s):  
Gastric Mucosa ◽  
Guinea Pig ◽  
Basal Lamina ◽  
Basal Cell ◽  
Morphological Changes ◽  
Short Circuit ◽  
Secretory Activity ◽  
Functional Changes ◽  
Ussing Chambers

Relationships between morphological and electrophysiological changes with low concentrations of ethanol on in vitro guinea pig gastric mucosa were investigated. Tissues mounted in Ussing chambers allowed recording of transepithelial potential difference (PD), resistance (R), short-circuit current (Isc), and acid secretion (H+). At selected times the mucosae were processed for morphological analysis. With luminal 10% ethanol there was a decrease in PD, R, Isc, and H+ within 1 min, and they eventually went to low steady-state values between 10 and 40 min. At 1 min many surface epithelial cells lifted off from the basal lamina but were still anchored by thin basal cell processes. After 10 min in ethanol many surface cells had completely detached from the basal lamina but remained connected to adjacent cells by their junctions. Numerous cytoplasmic blebs formed on both apical and basal cell surfaces. Concurrently, there was a significant increase in microvillus length. After 40 min most of the surface cells were detached from the basal lamina as sheets forming epithelial blisters. Upon ethanol washout there was epithelial cell reattachment to the basal lamina and a return of the PD, R, and Isc to control values within 40 min. Incubation of the luminal surface with 10% ethanol for 5 h resulted in a gradual rise of the PD, R, Isc, and H+ to control values by 4 h with a coincident return of the normal mucosal morphology. These studies indicate that ethanol has reversible and possibly adaptable effects on the in vitro guinea pig gastric mucosa and that the morphological changes are closely correlated with the decline and recovery of the electrical and secretory activity of the tissue.

Aldosterone stimulates K secretion prior to onset of Na absorption in guinea pig distal colon

1994 ◽  
Vol 266 (2) ◽  
pp. C552-C558 ◽  
Author(s):  
D. R. Halm ◽  
S. T. Halm
Keyword(s):  
Guinea Pig ◽  
Short Circuit ◽  
Short Circuit Current ◽  
Distal Colon ◽  
Ussing Chambers ◽  
High Affinity Receptor ◽  
Additive Increase ◽  
K Secretion ◽  
Affinity Receptor

Distal colon from guinea pig was stimulated in vitro by aldosterone in Ussing chambers that allowed measurement of short-circuit current (Isc) and tissue conductance (Gt). The response to aldosterone was delayed by approximately 20 min and resulted in a negative Isc, consistent with K secretion. Approximately 1 h later the Isc began to increase and eventually became positive, consistent with subsequent stimulation of Na absorption. The Na-absorptive response could be inhibited by mucosal amiloride without altering the rate of K secretion. Similarly, K secretion could be inhibited by serosal bumetanide without altering Na absorption. In the presence of spironolactone, actinomycin D, or cycloheximide, aldosterone failed to stimulate both K secretion and Na absorption. A dose response to aldosterone provided an apparent Kd of 2.6 +/- 0.5 nM, consistent with a high-affinity receptor coupled to this secretory response. Stimulation by the K secretagogue epinephrine did not produce an additive increase in K secretion, suggesting that the same cell type responds to both aldosterone and epinephrine and that the protein induced by aldosterone was not one of the membrane proteins responsible for K secretion.

Ouabain-sensitive H+-K+ exchange mechanism in the apical membrane of guinea pig colon

1989 ◽  
Vol 256 (6) ◽  
pp. G979-G988 ◽  
Author(s):  
Y. Suzuki ◽  
K. Kaneko
Keyword(s):  
Guinea Pig ◽  
Apical Membrane ◽  
Short Circuit ◽  
Secretion Rate ◽  
Bathing Solution ◽  
K Uptake ◽  
Uptake Mechanism ◽  
Ussing Chambers ◽  
Mucosal Side ◽  
86Rb Influx

The K+ uptake process across the apical membrane was investigated in relation to the ouabain-sensitive H+ secretion and the active transepithelial K+ absorption in guinea pig distal colon. The isolated colon was mounted between Ussing chambers or everted and fixed over a polyethylene tube (86Rb uptake measurements). The H+ secretion rate was determined from changes in pH of the weak-buffered mucosal bathing solution. K+ and Rb+ in the mucosal bathing solution powerfully stimulated H+ secretion. Cs+ also activated H+ secretion, though less than K+ or Rb+, whereas Na+, Li+, or choline scarcely did so. The increase in K+ concentration on the mucosal side reduced the degree of inhibition of H+ secretion by mucosal ouabain. 86Rb influx across the apical membrane (5.4 mM Rb+ in mucosal solution) was inhibited by approximately 90% by mucosal ouabain with the half-maximal effective dose being 3.9 +/- 1.0 X 10(-6) M; the half-maximal dose is not different from that for the inhibition of H+ secretion under similar conditions. The ouabain-sensitive component of 86Rb influx and the ouabain-sensitive H+ secretion rate were closely correlated when these rates were altered with either changes in the mucosal Rb+ concentration or addition of inhibitors such as acetazolamide, indomethacin, or 2,4-dinitrophenol. In the presence of K+ on the mucosal side, the ouabain-sensitive 86Rb influx was significantly reduced. Finally, the active transepithelial 42K absorption under short-circuit conditions was inhibited by approximately 50% with 4 X 10(-6) M and almost abolished with 10(-4) M ouabain on the mucosal side. It is concluded that the ouabain-sensitive K+ (Rb+) uptake mechanism, which is closely associated with H+ secretion, is present in the apical membrane of the colonic epithelial cells; the molecular mechanism of this uptake may be a H+-K+-ATPase. This K+ uptake is likely to mediate the apical membrane step of active transepithelial K+ absorption.

Stimulation of Cl- and HCO3- secretion by intramural cholinergic neurons in guinea pig antrum in vitro

1993 ◽  
Vol 264 (1) ◽  
pp. G118-G125 ◽  
Author(s):  
A. G. Suzuki ◽  
J. Kameyama ◽  
M. Tsukamoto ◽  
K. Kaneko ◽  
Y. Suzuki
Keyword(s):  
Guinea Pig ◽  
Short Circuit ◽  
Cholinergic Neurons ◽  
Short Circuit Current ◽  
Muscarinic Agonist ◽  
Serosal Side ◽  
Ussing Chambers ◽  
Bethanechol Chloride ◽  
Stimulation Of

Regulation of Cl- and HCO3- secretion by intramural cholinergic neurons was investigated in guinea pig antrum in vitro. Sheet preparations composed of the mucosa and the submucosa were mounted between Ussing chambers and bathed with buffer-free solution on the luminal surface and with HCO3(-)-CO2 solution on the serosal side. Short-circuit current (Isc), unidirectional fluxes of 36Cl and 22Na, and the luminal alkalinization rate (JOHSL) were determined. Electrical stimulation of the preparations elicited increases in both JOHSL and Isc, which were inhibited by tetrodotoxin (TTX) and atropine. Physostigmine also evoked TTX- and atropine-sensitive increases in JOHSL and Isc. Similar increases in JOHSL and Isc were observed when the muscarinic agonist bethanechol chloride (BCh) was added to the serosal side. The responses to BCh were not affected by TTX. The BCh-induced increase in JOHSL was largely abolished by removal of HCO3- or Na+ and addition of ouabain (serosal side) but was neither sensitive to Cl- removal nor associated with 22Na secretion. The increase in Isc induced by BCh was associated with the increase in 36Cl secretion and was inhibited by removal of Cl- or Na+ and by addition of bumetanide or ouabain (both, serosal side). These results suggest that the submucosal cholinergic neurons are involved via muscarinic receptors in the stimulation of epithelial HCO3- and Cl- secretion. For both HCO3- and Cl-, the cellular and membrane mechanisms of secretion induced by muscarinic stimulation, although not entirely clear, appear to be different from those occurring under baseline conditions.

scholarly journals β-Adrenergic activation of electrogenic K+ and Cl− secretion in guinea pig distal colonic epithelium proceeds via separate cAMP signaling pathways

2010 ◽  
Vol 299 (1) ◽  
pp. G81-G95 ◽  
Author(s):  
Susan T. Halm ◽  
Jin Zhang ◽  
Dan R. Halm
Keyword(s):  
Guinea Pig ◽  
Signaling Pathways ◽  
Time Course ◽  
Peptide Yy ◽  
Short Circuit ◽  
Short Circuit Current ◽  
Nucleotide Exchange ◽  
Adrenergic Stimulation ◽  
Camp Content ◽  
Adrenergic Activation

Adrenergic stimulation of isolated guinea pig distal colonic mucosa produced transient Cl− and sustained K+ secretion. Transient short-circuit current ( Isc) depended on β2-adrenergic receptors (β2-AdrR), and sustained Isc relies on a β1-AdrR/β2-AdrR complex. Epinephrine (epi) increased cAMP content with a biphasic time course similar to changes in epi-activated Isc (epi Isc). Inhibition of transmembrane adenylyl cyclases (tmACs) reduced peak epi Isc and cAMP to near zero without decreasing sustained epi Isc, consistent with cAMP from tmAC signaling for only Cl− secretion. Inhibition of soluble adenylyl cyclase (sAC) reduced sustained epi Isc and cAMP to near zero without decreasing peak epi Isc or cAMP, consistent with cAMP from sAC signaling for K+ secretion. Sensitivity to phosphodiesterase (PDE) inhibitors and peptide YY (PYY) stimulation further supported separate signaling for the two components. PDE3 or PDE4 inhibitors enhanced peak epi Isc but not sustained epi Isc, consistent with these PDEs as part of the β2-AdrR signaling domain. PYY suppressed peak epi Isc in a pertussis toxin (PTx)-sensitive manner, supporting Gαi-dependent inhibition of tmACs producing cAMP for Cl− secretion. Since PYY or PTx did not alter sustained epi Isc, signaling for K+ secretion occurred via a Gαi-independent mechanism. Presence of multiple sAC variants in colonic epithelial cells was supported by domain-specific antibodies. Responses to specific activators and inhibitors suggested that protein kinase A was not involved in activating peak or sustained components of epi Isc, but the cAMP-dependent guanine nucleotide exchange factor, Epac, may contribute. Thus β-adrenergic activation of electrogenic Cl− and K+ secretion, respectively, required tmAC- and sAC-dependent signaling pathways.

Morphology and electrophysiology of guinea pig gastric mucosal repair in vitro

1983 ◽  
Vol 244 (2) ◽  
pp. G171-G182 ◽  
Author(s):  
M. J. Rutten ◽  
S. Ito
Keyword(s):  
Steady State ◽  
Guinea Pig ◽  
Morphological Analysis ◽  
Short Circuit ◽  
Isotonic Saline ◽  
Short Circuit Current ◽  
Electrical Parameters ◽  
Mucosal Repair ◽  
Ussing Chambers

Guinea pig gastric mucosae stripped of their outer muscle layers were studied in Ussing chambers for up to 14 h. Ten minutes after the mucosae were mounted in the chamber, the electrical parameters were low but continued to rise over 90 min until steady-state potential difference (PD), resistance (R), and short-circuit current (Isc) were recorded. Morphological analysis during the first 10 min of the tissue in the chamber revealed gaps in the epithelium due to damaged cells. However, tissues examined after 20 min in the chamber showed little evidence of epithelial discontinuity. Thereafter, the initial rise in the electrical parameters was noted. After steady-state attainment, the lumen was exposed to 1.25 M NaCl for 5 min and then changed back to 150 mM NaCl. Ten minutes after washout and return to control solutions, the PD, R, and Isc had fallen to low values. At 30 min after washout of the NaCl, the PD, R, and Isc began to increase and after 2 h were back to control values. Morphological analysis of mucosae fixed up to 10 min after exposure to 1.25 M NaCl showed extensive damage and exfoliation of surface cells. However, by 30 min the epithelium was restored and had very few discontinuities, which was then followed by the return of the electrical parameters. The conclusions from these studies are 1) guinea pig gastric mucosae exposed to hypertonic NaCl on the luminal side will primarily result in surface epithelial cell destruction with an immediate drop in the transepithelial electrical values; 2) after return to isotonic saline the damaged mucosa can repair itself within minutes, which then allows the reestablishment of the transepithelial electrical parameters by 2 h; and 3) the good viability and reproducibility of this preparation present a suitable mammalian model system for the study of factors of mucosal repair.

Active potassium secretion by rabbit proximal colon

1986 ◽  
Vol 250 (4) ◽  
pp. G475-G483 ◽  
Author(s):  
S. K. Sullivan ◽  
P. L. Smith
Keyword(s):  
Short Circuit ◽  
Apical Cell ◽  
Distal Colon ◽  
Proximal Colon ◽  
Bathing Solution ◽  
Uptake Mechanism ◽  
Apical Cell Membrane ◽  
Ussing Chambers ◽  
K Concentration

Fluxes of K from mucosa to serosa or serosa to mucosa have been examined in stripped preparations of rabbit proximal and distal colon in vitro under short-circuit conditions in Ussing chambers. Results from these studies demonstrate that steady-state radioisotopic fluxes of K are achieved after 90 min and remain constant for at least 2 h. Determination of the K concentration dependence of the serosal-to-mucosal K flux revealed that this flux contains both saturable and nonsaturable components. Addition of ouabain (0.1 mM) abolished the saturable component of the serosal-to-mucosal K flux. The mucosal-to-serosal K flux is a linear function of K concentration between 1 and 20 mM under basal conditions. In paired tissues, serosal-to-mucosal K flux is always greater than mucosal-to-serosal flux under basal conditions resulting in net K secretion. However, addition of barium (2 mM) to the mucosal or serosal bathing solution had no significant effect on either unidirectional or net K fluxes. In addition, mucosal bumetanide (0.1 mM) or removal of Cl from both bathing solutions had no significant effect on unidirectional or net K fluxes. In rabbit distal colon, Cl removal from the bathing solutions significantly reduced serosal-to-mucosal K flux, resulting in net K absorption. These results indicate that rabbit proximal colon like rabbit distal colon actively secretes K. However, unlike distal colon the proximal colon does not possess an active K uptake mechanism at the apical cell membrane.(ABSTRACT TRUNCATED AT 250 WORDS)

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