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J A Schafer

Publications and source records attributed to J A Schafer.

At least 19 recordsLinked to original sources
PubMed1994

Sodium and water transport in cortical collecting duct of Dahl salt-resistant rat.

Studies were conducted to determine whether the cortical collecting duct (CCD) of the Dahl salt-resistant rat (inbred Rapp strain; R/Jr) exhibits the same responses to deoxycorticosterone (DOC; 2.5 mg as a depot injection in vivo, 3-8 days before experimentation) and arginine vasopressin (AVP, 220 pM in vitro) as the Sprague-Dawley (SD) [L. Chen, S.K. Williams, and J.A. Schafer. Am. J. Physiol. 259 (Renal Fluid Electrolyte Physiol. 28): F147-F156, 1990] and Dahl salt-sensitive (inbred Rapp strain, S/Jr) [C.T. Hawk and J.A. Schafer. Am. J. Physiol. 260 (Renal Fluid Electrolyte Physiol. 29): F471-F478, 1991] CCD. Qualitatively, the R/Jr CCD responded as in the other two strains: AVP elevated the osmotic water permeability (Pf, micron/s) from 0 to approximately 1,200; either AVP or DOC, when used alone, increased the lumen-to-bath 22Na+ flux (Jl-->b, pmol.min-1.mm-1) from the control range of 20-25 to approximately 40 and hyperpolarized the transepithelial voltage. AVP and DOC effects were synergistic, elevating Jl-->b to 90 +/- 5 (mean +/- SE) with both hormones, but this value was significantly lower than observed previously in both the SD and the S/Jr CCD, 125 +/- 6 and 140 +/- 6, respectively. However, bath-to-lumen fluxes (Jb--l) were also significantly lower than observed in the SD and S/Jr CCD. Because net fluxes (Jnet) in these experiments can be determined only as nonpaired differences between unidirectional fluxes, it is uncertain whether Jnet values in the R/Jr CCD are significantly lower than in the SD or S/Jr CCD.(ABSTRACT TRUNCATED AT 250 WORDS)

Amiloride↗
PubMed1993

Clonidine, but not bradykinin or ANP, inhibits Na+ and water transport in Dahl SS rat CCD.

We examined the actions of potentially natriuretic autacoids in the isolated perfused cortical collecting duct (CCD) dissected from inbred Dahl (Rapp strain) salt-sensitive rats (SS). Atrial natriuretic peptide (ANP, 10 nM), bradykinin (BK, 10 nM), and clonidine (1 microM) were studied to determine their effects on the lumen-to-bath flux of 22Na+ (J1-->b, pmol min-1 mm-1), hydraulic conductivity (Pf, micron/s), and transepithelial voltage (VT, mV). ANP and BK have been shown by others to significantly reduce net Na+ reabsorption and hydraulic conductivity in the Sprague-Dawley (SD) rat CCD, but previous results from our laboratory showed no ANP or BK effect in the SD CCD. In the present study, we were also unable to observe any effect of either ANP or BK in the SS rat CCD. However, in the presence of AVP, clonidine (a partial alpha 2-adrenergic receptor agonist) significantly reduced J1-->b and Pf from 139 +/- 6 (SEM) to 88 +/- 7 and from 959 +/- 176 to 490 +/- 73, respectively. In addition, clonidine significantly depolarized VT from -14.5 +/- 2.8 to -11.2 +/- 1.8. However, unlike its effects in the SD rat CCD, yohimbine (300 nM, an alpha 2-adrenergic receptor antagonist) did not significantly reverse the effects of clonidine on J1-->b, Pf or VT in the SS rat CCD.

Animals↗
PubMed1993

Inhibition by epinephrine of AVP- and cAMP-stimulated Na+ and water transport in Dahl rat CCD.

We examined the effects of epinephrine in perfused cortical collecting ducts (CCD) isolated from inbred Dahl-Rapp salt-sensitive (SS) and salt-resistant (SR) rats and from Sprague-Dawley (SD) rats. Rats were treated with 2.5 mg deoxycorticosterone pivalate (DOC; depot injection 4-9 days before study), and the CCD were treated with 220 pM vasopressin (AVP) to maximize Na+ transport. In CCD from all three strains 10 microM epinephrine in the bathing solution completely inhibited net Na+ transport, osmotic water permeability (Pf), and transepithelial voltage. In the SS CCD, epinephrine increased the fractional resistance of the luminal membrane to the same extent as 10 microM amiloride, indicating that it blocked the amiloride-sensitive conductance of the luminal membrane. Even at 100 nM epinephrine inhibited 80-100% of Na+ and water transport, and 1 microM yohimbine reversed or prevented these effects. In SS CCD, 0.1 mM 8-bromoadenosine 3',5'-cyclic monophosphate (8-BrcAMP) plus 0.1 mM 3-isobutyl-1-methylxanthine in place of AVP increased lumen-to-bath Na+ flux (J1-->b) from 56 +/- 5 to 143 +/- 3 pmol.min-1 x mm-1 and Pf from 6 +/- 12 to 1067 +/- 152 microns/s, but 100 mM epinephrine still significantly inhibited cAMP-stimulated J1-->b and Pf by 40 +/- 5% and 31 +/- 9%, respectively. Similar results were observed in the SR and SD rat CCD; however, the ability of yohimbine to reverse the epinephrine effect on cAMP-dependent transport was variable among the rat strains. We conclude that epinephrine acts via an alpha 2-receptor to inhibit adenylate cyclase but that at least one additional intracellular second messenger system may be involved.

Adrenergic alpha-Antagonists↗
PubMed1993

Intracellular Ca2+ and PKC activation do not inhibit Na+ and water transport in rat CCD.

Experiments examined the effects of elevation of intracellular calcium concentration ([Ca2+]i) or activation of protein kinase C (PKC) on Na+ and water transport in the rat cortical collecting duct (CCD). We measured the lumen-to-bath 22Na+ flux (J1-->b), transepithelial voltage (VT), and water permeability (Pf) in CCD from deoxycorticosterone (DOC)-treated rats. Ionomycin (0.5 and 1 microM) and thapsigargin (1 and 2 microM) were used to increase [Ca2+]i. Phorbol 12-myristate 13-acetate (PMA; 0.3 and 1 microM) and oleoyl-acetyl-glycerol (OAG; 100 microM) were used as activators of PKC. [Ca2+]i was measured in isolated perfused tubules using the fluorescent dye fura 2. When added to the bathing solution, 220 pM arginine vasopressin (AVP) failed to affect [Ca2+]i, whereas 1 microM ionomycin increased [Ca2+]i by 103 +/- 15% and 2 microM thapsigargin increased [Ca2+]i by 24 +/- 4%. In flux studies, neither ionomycin nor thapsigargin affected J1-->b or Pf, although ionomycin caused marked morphological changes. Ionomycin also failed to alter either parameter in tubules from non-DOC-treated rats. Neither 100 microM OAG nor 1 microM PMA affected J1-->b or Pf. OAG at 50 microM had no effect on VT or transepithelial resistance, indicating no inhibition of conductive Na+ transport. We conclude that increased [Ca2+]i and PKC activation do not affect J1--b or Pf in the rat CCD. These findings may account for the sustained increase in J1--b produced in the rat CCD by AVP.

Animals↗
PubMed1992

Regulation of Na+ channels in the cortical collecting duct by AVP and mineralocorticoids.

A variety of experimental approaches have shown that AVP and mineralocorticoids stimulate Na+ transport through their effects on the number and kinetic properties of amiloride-sensitive Na+ channels in the apical membrane. The different mechanisms by which AVP and mineralocorticoid act on the Na+ channel provide a basis for synergism in their actions, perhaps by a scheme such as that proposed in Figure 5. However, the details of this interaction will require a better understanding of the molecular details involved in activating quiescent channels, increasing their open probability, and reorientating or inserting channels to an operational position in the apical membrane. Electrophysiological and biochemical approaches have gone a long way toward elucidating some of these molecular details. But the latter approach in particular has indicated that the Na+ channel may have multiple regulatory subunits and thus be a target for several intracellular second messengers and autacoids other than those involved in the actions of AVP and aldosterone. The challenges for future research in this area are multiple. It seems likely that the primary amino acid sequence of the channel subunits will soon become available from cloning and sequencing approaches, but the application of this knowledge to understanding how the subunits are integrated into the complete protein and mediate regulatory signals will be a formidable task. It will be important to determine the normal extracellular signals (other than aldosterone and AVP) and the associated intracellular second messengers that alter channel activity. It will also be important to understand how some species such as the rabbit may "turn off" the stimulatory effect of AVP on Na+ reabsorption in the CCD, and how this regulatory process is altered when these cells are cultured. At the whole animal level, it will also be important to investigate whether changes in one or more of the normal regulatory pathways that impinge on the Na+ channel might be involved in a diminished ability to excrete a salt load, as is observed in some models of hypertension. All of these issues need to be understood at the molecular level, and it seems likely they will provide exciting physiological insights at all levels.

Aldosterone↗
PubMed1992

AVP stimulates Na+ transport in primary cultures of rabbit cortical collecting duct cells.

Immunodissected rabbit cortical collecting duct (CCD) cells were grown in primary culture on permeable membrane supports. Transepithelial voltage, Na+, K+, and H+ gradients developed as expected for a mixed population of principal and intercalated cells. The amiloride-sensitive short-circuit current (Isc) was measured in Ussing chambers as an index of Na+ transport via apical membrane Na+ channels. Treatment of the cells in culture with 10 nM aldosterone for 48 h increased Isc from 7.4 +/- 1.4 to 19.3 +/- 3.2 microA/cm2. In contrast to the native rabbit CCD, 220 pM arginine vasopressin (AVP) produced a rapid and stable (greater than 60 min) increase in Isc to 15.8 +/- 2.0 and 29.0 +/- 3.8 microA/cm2 in untreated and aldosterone-treated cultures, respectively. Although prostaglandin E2 (PGE2) inhibits Na+ transport in the native rabbit CCD, it did not in the cultured cells, and it has previously been shown that PGE2 inhibition of AVP-dependent adenosine 3',5'-cyclic monophosphate production is lost in culture (W. K. Sonnenburg and W. L. Smith, J. Biol. Chem. 263: 6155-6160, 1988). We conclude that the development of a stable stimulation of Na+ transport by AVP is linked to the loss of the inhibitory effects of PGE2.

Aldosterone↗
PubMed1991

Structural changes induced by osmotic water flow in rabbit proximal tubule.

When a transepithelial osmotic difference was imposed in perfused proximal straight tubules (270 mOsm/kg H2O in the lumen and 290 in the bath) in the absence of bath colloid, a severe vacuolation (appearance of lucent spaces) developed within the epithelium such that view of the lumen border was obscured within 5 +/- 1 min (N = 13 tubules at 23 degrees C). This vacuolation was less severe if the bath was hypotonic to the lumen or if the magnitude of the osmotic difference was reduced. If colloid (6% wt/vol of either bovine serum albumin or 70,000 molecular wt dextran) was included in the bathing medium, vacuolation was either not observed or was minimal, but became severe upon removal of the colloid and obscured the lumen within 6 +/- 1 min (N = 8 for albumin and N = 4 for dextran at 23 degrees C). At 38 degrees C, vacuolation obscured the lumen within 4 +/- 1 min following the removal of albumin (N = 5). ANOVA suggests that none of the times for vacuolation to occur differed. The rate of passive volume flow due to the osmotic difference was unaffected by vacuolation (0.9 +/- 0.1 nl.min-1.mm-1 with albumin to 0.8 +/- 0.1 without albumin and vacuolated, N = 8 at 23 degrees C, P greater than 0.2 using a paired t-test). Electron microscopic examination of tubules fixed after vacuolation showed lucent spaces within the cytoplasm. These results suggest that the presence of serosal colloid protected the epithelial cells from injury during rapid transepithelial water flow. The mechanism for this protective effect is not apparent, but may be related to effects of colloid in maintaining normal volume absorption in the proximal nephron.

Animals↗
PubMed1991

Na+ transport in isolated rat CCD: effects of bradykinin, ANP, clonidine, and hydrochlorothiazide.

We examined the effects of bradykinin (BK), atrial natriuretic peptide (ANP), hydrochlorothiazide (HCTZ), and clonidine on Na+ transport in isolated perfused cortical collecting ducts from rats treated with deoxycorticosterone. Arginine vasopressin was present in the bathing solution at 220 pM. Clonidine (1 microM, bathing solution) depolarized transepithelial potential difference (PDT) from -11.9 +/- 2.0 (SE) to -7.4 +/- 1.7 mV (P less than 0.001), hyperpolarized basolateral membrane potential difference (PDbl) from -85 +/- 1 to -87 +/- 1 mV (P less than 0.01), and increased the fractional resistance of the apical membrane (FRa) from 0.81 +/- 0.02 to 0.86 +/- 0.02 (P less than 0.03), indicating that it inhibited the Na+ conductance of the luminal membrane. BK (1 or 10 nM) or ANP (10 nM) in the bathing solution had no effect on PDT, PDbl, or FRa. BK, ANP, or 0.1 mM luminal HCTZ also had no effect on lumen-to-bath 22Na+ flux (J1----b), whereas we showed previously that clonidine inhibits J1----b by 30% (L. Chen, M. Paris, S. K. Williams, M. C. Reif, and J. A. Schafer. Kidney Int. 37: 366, 1990). Luminal addition of Na+ channel blockers amiloride (10 microM) or benzamil (1 microM) reduced J1----b to a level not significantly different from bath-to-lumen 22Na+ flux measured previously (M. Reif, S. L. Troutman, and J. A. Schafer. J. Clin. Invest. 77: 1291-1298, 1986), and neither BK nor HCTZ had any further effect. These results show that transcellular Na+ transport occurs exclusively through the apical membrane amiloride-sensitive channel, and this conductance is inhibited by clonidine but not by BK, ANP, or HCTZ.

Amiloride↗
PubMed1991

Effects of AVP and deoxycorticosterone on Na+ and water transport in the Dahl salt-sensitive rat CCD.

Cortical collecting ducts (CCD) from inbred Dahl salt-sensitive rats were perfused in vitro to study effects of arginine vasopressin (AVP, present in the bath) and deoxycorticosterone pivalate (DOC) pretreatment on lumen-to-bath and bath-to-lumen fluxes of 22Na+ (J1----b and Jb----1 in pmol.min-1.mm-1, respectively), hydraulic conductivity (Pf, microns/s), and transepithelial voltage (VT, mV). J1----b was 37.1 +/- 5.3 (mean +/- SE) in untreated rats and increased to 83.2 +/- 15.9 with AVP. VT increased from -0.3 +/- 0.6 to -7.0 +/- 2.0. In DOC-pretreated rat CCDs, baseline J1----b was higher (85.1 +/- 7.6) as was VT (-11.3 +/- 1.7); J1b----b and VT were doubled with AVP addition (185.6 +/- 18.6 and -21.7 +/- 2.3, respectively). Thus J1----b in AVP-stimulated CCDs from untreated rats was not significantly different from control (no AVP) J1----b from DOC-pretreated rats; however, AVP produced a greater J1----b increase in the latter CCDs. Neither AVP nor DOC had an effect on Jb----1, which ranged from 25 to 50. Benzamil reduced J1----b to values not significantly different from Jb----1, and VT became zero in CCDs treated with both AVP and DOC, indicating Na+ transport stimulated by both hormones occurs through apical membrane Na+ channels. Pf increased from 59 +/- 80 to 1,072 +/- 176 with AVP addition to untreated rat CCDs and was unaltered by DOC or benzamil. Thus the Dahl rat CCD exhibits a transport response to AVP and DOC that is indistinguishable from that observed in Sprague-Dawley rats, as previously reported by this laboratory.

Absorption↗
PubMed1991

Clonidine and PGE2 have different effects on Na+ and water transport in rat and rabbit CCD.

We examined the effects of prostaglandin E2 (PGE2) and the adrenergic agonist clonidine on osmotic water permeability (Pf), lumen-to-bath 22Na+ flux (Jl----b), and transepithelial voltage (VT) in isolated perfused cortical collecting ducts (CCD) from rats and rabbits. Although PGE2 inhibited arginine vasopressin (AVP)-dependent Jl----b and Pf in CCDs from deoxycorticosterone (DOC)-treated and untreated rabbits, 0.1-10 microM PGE2 had no inhibitory effect on any of these transport parameters in CCDs from DOC-treated rats in presence of AVP. On the other hand, clonidine (1 microM in bathing solution) reversibly inhibited AVP-dependent Pf, Jl----b, and VT in the rat CCD by 30-40%, and 0.3 microM yohimbine, a specific alpha 2-adrenoceptor antagonist, reversed these effects. However, we were unable to demonstrate any inhibitory effect of 1-10 microM clonidine on Pf, Jl----b, or VT in the rabbit CCD using a variety of protocols. These results are consistent with the pattern of inhibition of AVP-dependent adenosine 3',5'-cyclic monophosphate (cAMP) production in the rat and rabbit CCD in that PGE2 inhibits both transport and cAMP production in the rabbit but not the rat CCD, and clonidine inhibits both transport and cAMP production in the rat but not the rabbit CCD [D. Chabardès, C. Brick-Ghannam, M. Montégut, and S. Siaume-Perez, Am. J. Physiol. 255 (Renal Fluid Electrolyte Physiol. 24): F43-F48, 1988].

Animals↗
PubMed1990

Principal cells of cortical collecting ducts of the rat are not a route of transepithelial Cl- transport.

The rat cortical collecting duct (CCD) exhibits high rates of NaCl reabsorption when stimulated by mineralocorticoid and antidiuretic hormone (ADH). The present study was undertaken to determine if there is significant transcellular Cl- movement across the principal cells of the rat CCD. CCDs were dissected from kidneys of rats that had been injected with deoxycorticosterone (5 mg, i.m.) 2-9 days prior to the experiment. The ducts were perfused in vitro with identical perfusing and bathing solutions, except that 200 pmol.l-1 ADH was added to the bathing solutions. The basolateral membrane voltage (PDbl) of principal cells was -77 +/- 1 mV and the luminal membrane voltage (PD1) was -68 +/- 1 mV (mean +/- SEM, n = 124). Separate impalements with single-barrelled Cl(-)-selective microelectrodes gave an apparent intracellular Cl- activity of principal cells of 17 +/- 2 mmol.l-1. Transepithelial PD and PDbl were unaffected by luminal furosemide, hydrochlorothiazide (HCT), 4-acetamido-4-isothiocyanostilbene2,2-disulphonic acid, (SITS), or the Cl- channel blocker 5-nitro-2-(3-phenylpropylamino)-benzoic acid (NPPB); bath addition of SITS or the Cl- channel blocker diphenylamino-2-carboxylic acid; or replacement of bath HCO3- by Cl-. The intracellular Cl- activity (a(cell)Cl) also remained unchanged with the addition of HCT, SITS or the Cl- channel blockers to either the perfusing or bathing solutions, or with replacement of the bathing solution HCO3-.(ABSTRACT TRUNCATED AT 250 WORDS)

Absorption↗
PubMed1990

Transepithelial osmolality differences, hydraulic conductivities, and volume absorption in the proximal tubule.

The weight of current evidence indicates that the proximal tubule has a high transepithelial osmotic water permeability in the range of 3500-6000 microns/sec, which is attributable in large part to the high water permeabilities of the cell membranes. Water movement through these membranes may occur through specialized, proteinaceous channels that can be blocked by sulfhydryl reagents. The water channels probably exclude even the smallest solutes and allow only single file movement of the water molecules as do the water channels previously described in the red blood cell and vasopressin-responsive epithelia. If a significant fraction of the water flow also occurs through the junctional complexes, it seems likely that these junctions could be a site for solute solvent coupling which would contribute to solute absorption by solvent drag and which would be responsible for non-unity reflection coefficients for some solutes such as Na+ and Cl-. This possibility is still a matter of vigorous debate. Since the transepithelial water permeability is high, only a very small osmolality difference (1-10 mOsM) is required to drive normally observed rates of volume absorption both in vivo and in vitro. The osmolality difference is produced at least in part by dilution of the luminal fluid and is possibly augmented by the development of interstitial hyperosmolality because of the rapid transport of preferentially absorbed solutes. In the future it is likely that the most important work in this field will relate to the factors that alter transepithelial water permeability and the solute and water permeabilities of the junctional complexes. Investigation in this area is essential in understanding how changes in capillary and interstitial hydrostatic and colloid osmotic pressure may affect volume absorption.

Absorption↗
PubMed1990

Vasopressin and mineralocorticoid increase apical membrane driving force for K+ secretion in rat CCD.

Cortical collecting ducts (CCD) from untreated Sprague-Dawley rats were perfused and bathed in vitro with modified Krebs-Ringer solutions. Arginine vasopressin (AVP;100 microU/ml) in the bathing solution hyperpolarized the transepithelial voltage (PDT, mV) from -2.3 +/- 0.7 (control) to -6.0 +/- 1.1 (n = 22) and decreased the transepithelial resistance from 64 +/- 7 to 54 +/- 7 omega.cm2 (n = 21). AVP depolarized the basolateral membrane voltage of principal cells (PDbl) only slightly (but significantly by paired statistical comparison) from -85 +/- 1 to -84 +/- 1 mV (n = 9), with a fall in the fractional resistance of the apical membrane (FRa) from 0.82 +/- 0.03 to 0.77 +/- 0.05 (n = 9). Luminal amiloride (10 microM) produced no change in FRa in the absence of AVP, but in the presence of AVP increased FRa to the same level observed in the absence of AVP. The changes with AVP were significantly less than those observed by us previously in deoxycorticosterone (DOC)-treated animals (E. Schlatter and J. A. Schafer. Pfluegers Arch. 409:81-92, 1987), indicating that the observed synergism between DOC and AVP in stimulating Na+ absorption is attributable to a greater increase in the Na+ conductance in the apical membrane of principal cells with AVP in the DOC-treated CCD than in the normal. Furthermore, we have calculated that the depolarization of apical membrane voltage resulting from the increased Na+ conductance produced by either or both AVP and DOC increases the driving force for K+ exit across the apical membrane in proportion to the previously measured increase in secretion. This increase in driving force may be sufficient to explain the increased K+ secretion produced by these hormones with no change in the apical membrane K+ conductance.

Amiloride↗
PubMed1990

Differences in synergistic actions of vasopressin and deoxycorticosterone in rat and rabbit CCD.

We examined the time course of changes in osmotic water permeability (Pf, micron/s), lumen-to-bath 22Na+ flux (Jl----b, pmol.min-1.mm-1), and transepithelial voltage (VT, mV) in response to arginine vasopressin (AVP) in isolated perfused rat and rabbit cortical collecting ducts (CCD). CCDs were isolated from "normal" (untreated) animals and animals 4-14 days after an intramuscular depot injection of deoxycorticosterone pivalate (DOC; 35 mg/rabbit, 5 mg/rat). In the normal rat CCD, 220 pM AVP added to the bathing solution increased Pf from approximately 0 to 981 +/- 120, Jl----b from 24 +/- 5 to 62 +/- 9, and VT from -0.7 +/- 0.6 to -4.0 +/- 0.6 (mean +/- SE; P less than 0.05). The effect of AVP on Pf in DOC-treated rats was not significantly different, but the effects on Jl----b and VT were greater. Jl----b rose from a control value of 78 +/- 16 to 192 +/- 14 with AVP, and VT hyperpolarized from -4.2 +/- 1.6 to -12.2 +/- 0.4. All of these effects of AVP in the normal and DOC-treated rat CCD persisted for greater than or equal to 90-150 min. In DOC-treated rabbit tubules, AVP caused a stable increase in Pf from 9 +/- 6 to 800 +/- 199, which was significantly greater than in normal tubules (379 +/- 32). In the normal rabbit CCD we confirmed previous observations that AVP produces only a transient increase in Na+ absorption and VT; Jl----b increased from 76 +/- 4 to 96 +/- 7 during the period 15-25 min after AVP addition but subsequently fell to 80 +/- 6; VT transiently hyperpolarized from -55 +/- 7 to -64 +/- 8 at 3-8 min after AVP and then fell to -54 +/- 9. In DOC-treated rabbit CCDs, AVP produced no change whatsoever in Jl----b, which was not significantly different from the control value of 99 +/- 7, nor in VT, which was -68 +/- 8 mV. These results indicate fundamental differences in the response of the rat and rabbit CCD to AVP and the presence or absence of synergistic interactions with mineralocorticoids, which may relate to their differing response to other autacoids.

Animals↗
PubMed1990

cAMP mediates the increase in apical membrane Na+ conductance produced in rat CCD by vasopressin.

Experiments were conducted to determine if adenosine 3',5'-cyclic monophosphate (cAMP) mediates the stimulation of Na+ absorption by arginine vasopressin (AVP) in isolated perfused cortical collecting ducts (CCD) from rats treated with deoxycorticosterone pivalate (5 mg im) 5-9 days before study. AVP (220 pM) in the bathing solution hyperpolarized the transepithelial voltage (PDT) from -4.0 +/- 0.8 (SE) to -15.1 +/- 1.4 mV (n = 9, P less than 0.001) and decreased the transepithelial resistance (RT) from 40 +/- 8 to 33 +/- 6 omega.cm2 (n = 5, P less than 0.025). Bath addition of 0.2 mM dibutyryl cAMP (DBcAMP), 0.1 mM isobutylmethylxanthine (IBMX), 0.1 mM DBcAMP plus 0.1 mM IBMX, and 10 or 50 microM forskolin produced the same effects, reversibly hyperpolarizing PDT by 7.0-11.5 mV and decreasing RT by 6-12 omega.cm2. Addition of 10 microM amiloride to the luminal perfusate reduced PDT from -0.9 to +2.0 mV and increased RT in the presence or absence of any of the test agents. Addition of DBcAMP + IBMX or 50 microM forskolin to the bathing solution also reversibly depolarized the basolateral membrane voltage of principal cells by 1-2 mV and decreased the apical membrane fractional resistance from 0.82-0.84 to 0.72-0.77. Both effects were reversed by addition of amiloride to the luminal perfusate. These results demonstrate that cAMP is the intracellular mediator of the increase in apical membrane Na+ conductance produced by AVP in the rat CCD.

1-Methyl-3-isobutylxanthine↗