Regulation of L-alanine transport systems A and ASC by cyclic AMP and calcium in a reptilian duodenal model
Laboratorio de Fisiología Animal, Departamento de Biología Animal, Universidad de La Laguna, 38206 Tenerife, Spain
* Author for correspondence (e-mail: madiaz{at}ull.es)
Accepted 4 February 2003
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Summary |
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Key words: system A, system ASC, neutral amino acid transport, L-alanine transport, intracellular transducers, cyclic AMP, calcium, Gallotia galloti
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Introduction |
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Pioneering work by Edmonson and Lumeng
(1980) on isolated rat
hepatocytes showed that glucagon stimulation of neutral amino acid transport
was biphasic, with short- and long-term effects. Although the first phase was
totally insensitive to inhibitors of protein and RNA synthesis, the second
phase was completely prevented by cycloheximide. Later studies have
demonstrated that the rapid stimulation of L-alanine transport by glucagon is
caused by the rapid generation of cyclic AMP (cAMP), and it has been suggested
that the short-term stimulation of alanine transport is secondarily due to a
cAMP-mediated membrane hyperpolarization
(Moule et al., 1987
). Although
this model of regulation has been generally accepted for hepatocytes, it has
not been validated for other preparations expressing similar transport
systems. In fact, studies performed in vascular endothelial cells
(Escobales et al., 1994
),
kidney cortex (Goldstone et al.,
1983
), lymphocytes (Woodlock
et al., 1989
), glioblastoma cells
(Zafra et al., 1994
) and
placental slices (Karl et al.,
1988
; Ramamoorthy et al.,
1992
) have pointed out that alanine transport is acutely regulated
by intracellular calcium or diacylglycerol but not by cAMP. Interestingly,
despite the existence of multiple distinguishable transport systems for
neutral amino acids within each cell type, the regulation of transport
activity appears to be restricted to individual systems. For instance, neutral
amino acid transport systems in the plasma membrane of hepatocytes is carried
out by systems A, ASC and L, but only system A appears to be subjected to
adaptive regulation and pancreatic hormones
(Gazzola et al., 1981
;
Handlogten and Kilberg, 1984
;
Shotwell et al., 1983
).
In the small intestine of vertebrates, neutral amino acid transport is
catalyzed by several Na+-dependent and Na+-independent
systems (for reviews, see Kilberg et al.,
1993; Munck and Munck,
1994
; Stevens,
1992
). Nonetheless, despite extensive description of amino acid
transport pathways, kinetic properties, energy requirements and membrane
localization in isolated epithelia, enterocytes and brush-border membranes
from different animal models, studies describing short-term regulation of
intestinal neutral amino acid transport across epithelial cells are scant.
The purpose of this study was to investigate the regulation of L-alanine
transport across the lizard duodenum by increased intracellular cAMP and
calcium levels. This epithelium has been studied over the past two decades,
and the different pathways for neutral amino acid transport have been recently
identified and characterised physiologically
(Díaz et al., 2000;
Medina et al., 2001
).
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Materials and methods |
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Animals and solutions
Adult male lizards (Gallotia galloti sauria, lacertidae) weighing
2540 g were sacrificed by spinal transection and the duodenum was
removed and rinsed in ice-cold bathing solution. The standard Ringer solution
contained 107 mmol l1 NaCl, 4.5 mmol l1
KCl, 25 mmol l1 NaHCO3, 1.8 mmol
l1 Na2HPO4, 0.2 mmol
l1 NaH2PO4, 1.25 mmol
l1 CaCl2 and 1.0 mmol l1
MgCl2 and had a final pH of 7.3. Duodenal segments were mounted in
water-jacketed Ussing chambers with an exposed area of 0.21 cm2 and
bathed on both sides with 4 ml of Ringer solution. Chambers were continuously
gassed with 5% CO2 and 95% O2 and the temperature was
maintained at 27°C. In some experiments, choline was used to replace
sodium ions in the bathing solutions.
Electrical measurements
The electrical measurements were made as described previously
(Gómez et al., 1986)
using calomel (for voltage sensing) and Ag/AgCl electrodes (for current
passage) connected to the bathing solutions through 4% (v/w) agar bridges.
Electrical measurements were continuously monitored with an automatic
computer-controlled voltage-clamp device (AC-microclamp) and hardcopied. The
tissues were first incubated under open-circuit conditions for 20 min and then
under short-circuit conditions; i.e. the potential difference (PD) and the
short-circuit current (Isc) were determined every minute.
Every 5 s, the tissues were pulsed with 1 s ± 10 µA pulses, and,
from the displacement of the PD, the tissue conductance
(Gt) was derived. Corrections for electrode offset
potential and solution resistance were determined throughout the experiments
and stored in the computer-controlled voltage-clamp device.
Transepithelial fluxes
Unidirectional amino acid fluxes were measured under short-circuit
conditions using the procedure described in detail by Diaz et al.
(2000). Briefly, 20 min after
the tissue was properly mounted in the chamber, 185 kBq of the appropriate
labelled substrate (2,3[H]L-alanine or
[14C]MeAIB) was added to the serosal or mucosal side of the tissue.
After an additional 20 min period, by which isotope fluxes had reached steady
state, duplicate 200 µl aliquot samples were taken from the unlabelled side
at regular 20 min intervals for 1 h and replaced by an equal volume of Ringer
solution. Inhibition experiments were carried out by adding small volumes (100
µl) of concentrated stock solution containing the amino acid or analogue to
the mucosal and/or serosal compartments. The unidirectional and net fluxes
were determined using a computer program written in our laboratory
(Díaz and Cozzi, 1991
),
which also provided the statistical tools required for data analysis.
Cyclic AMP determination
Tissue cAMP accumulation in response to forskolin and theophylline was
determined on freshly isolated tissues. Once removed, tissues were washed
several times in cold saline, weighed, minced into small pieces and
immediately transferred to the standard bathing solution. Vehicles and drugs
were added at time zero to control and test tubes, respectively, and allowed
to preincubate for different times. Samples were then homogenized in a
Tris/EDTA buffer (0.5 ml per 100 mg tissue mass) at pH 7.5. Samples were then
deproteinized by heat in boiling oil for 3 min and centrifuged at 1000
g for 5 min. Duplicated 50 µl samples from each supernatant
were taken for cAMP determination using a standard cAMP assay kit (Amersham
Ibérica).
Statistical analysis
Results are expressed as means ± S.E.M. Statistical comparisons of
mean values were made using two-tailed Student's t-test or one-way
analysis of variance (ANOVA) followed by Tukey's test, where appropriate. In
some experiments, PD and Isc values were normalized by
transformation to percentage of maximal change within each experiment.
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Results |
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Bioelectrical parameters were measured under short-circuit conditions for
the different compounds and the results are illustrated in
Table 1. The tissues were
maintained for 2040 min under open-circuit conditions and were then
short-circuited. As has been reported several times for this same preparation
(Gómez et al., 1986;
Lorenzo et al., 1989b), no differences were observed between the transmural
potential difference (PD), short-circuit current (Isc) and
tissue conductance (Gt) between stable open-circuit and
short-circuit measurements. As can be seen in
Table 1, under control
conditions, transepithelial PD was approximately 2.0 mV and the mean
Isc was 0.77 µequiv. cm2
h1, the serosal side of the tissue being electrically
positive compared with the luminal side, which parallels our previous
observations (Gómez et al.,
1986
; Medina et al.,
2001
). Both PD and Isc were significantly
increased by forskolin, theophylline and db-cAMP, in agreement with most
studies performed in different animal models, including lizard intestine
(Bridges et al., 1983
;
Díaz and Lorenzo, 1991
;
Lorenzo et al., 1989a). The tissue conductance remained unaltered throughout
the experiments at approximately 8 mS cm2.
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Effects of forskolin and theophylline on intracellular cAMP
concentration
In order to test whether the effects of forskolin and theophylline on
L-alanine transport across lizard duodenum could be attributed to changes in
the intracellular levels of cAMP, we performed radioimmunoassays to determine
the time-course of cAMP concentration in isolated mucosal preparations. The
results shown in Fig. 2 revealed that both forskolin (10 µmol l1) and
theophylline (3 mmol l1) significantly increased cAMP
levels, although the time-course and the magnitude of change were clearly
different for the two drugs. Thus, cAMP concentration reached a maximal value
of 31.87±5.1 pmol tube1 10 min after the exposure of
mucosal tissues to 10 µmol l1 forskolin. On the other
hand, theophylline induced a transient increase of cAMP level that was much
lower (8.8±0.5 pmol tube1) than for forskolin, this
value being reached 5 min after exposure to the secretagogue. In addition, the
effect of forskolin was maintained for the rest of the experiment whereas the
effect of theophylline rapidly declined to control values 20 min after drug
exposure.
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Effects of calcium ionophore A23187 and calcium-free solutions on
total L-alanine transport
In the next series of experiments, manoeuvres known to alter cellular
calcium homeostasis were tested for their ability to affect L-alanine
transport across the lizard duodenum. Initially, the effects of calcium
ionophore A23187 (0.5 µmol l1) on unidirectional and net
L-alanine fluxes were analysed. As can be observed in
Fig. 3A, addition of calcium
ionophore to both sides of the tissue brought about a considerable decrease in
Jnet, this effect being entirely attributable to the
reduction of Jms. The remaining Jnet
(2.76±1.19 nmol cm2 h1) was still
significantly different from zero (P<0.05, N=16),
suggesting that a fraction of L-alanine transport was unaffected by the
ionophore. Despite the dramatic reduction of L-alanine transport by A23187, PD
and Isc (but not Gt) were increased by
the ionophore (Table 2),
indicating that increased calcium levels had stimulated a conductive
process.
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In another set of experiments, the effects of extracellular calcium replacement on L-alanine fluxes and biolectrical parameters were determined. The results shown in Fig. 3B and Table 2 indicate that none of the parameters under study were modified by calcium replacement.
Effects of db-cAMP and A23187 on individual L-alanine transport
systems
We have previously demonstrated that, under short-circuit conditions, the
overall L-alanine transport in the lizard duodenum could be completely
explained by the simultaneous participation of two Na+-dependent
transport systems, endowed with properties of systems ASC and A, plus one
Na+-independent electrogenic carrier
(Díaz et al., 2000;
Medina et al., 2001
). These
three different transport systems could be individually dissected by means of
a strategy based on the differential substrate affinities and sodium
dependence. In order to determine whether individual L-alanine transport
systems were equally affected by cAMP and calcium, we performed experiments
aimed at assessing the effects of these intracellular transducers on each
individual transport system.
First, we measured L-alanine fluxes under short-circuit conditions using
Na+-free solutions. Under these conditions, active L-alanine
transport was carried by an electrogenic cycloleucine- and
2-aminobicyclo-(2,2,1)-heptane-2-carboxilic acid (BCH)-sensitive pathway
(Díaz et al., 2000). The
results shown in Figs 4A and
5A indicate that neither
db-cAMP nor A23187 affected unidirectional or net L-alanine fluxes, which
strongly suggests that changes in the activity of the
Na+-independent carrier were not responsible for the effects on the
overall L-alanine transport observed above, but rather these resulted from the
activities of the Na+-dependent systems.
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Therefore, we measured the effects of cAMP and calcium on the activity of
the two Na+-dependent transport pathways identified in the lizard
duodenum, i.e. system A and system ASC. Initially, system A activity was
determined by using the specific N-methylated derivative
-methylamino-isobutiric acid (MeAIB) in the presence of sodium. Our
results showed that neither unidirectional nor net MeAIB fluxes were altered
by db-cAMP at concentrations known to increase duodenal L-alanine transport
(Fig. 4B). Conversely, addition
of A23187 significantly increased Jms (but not
Jsm), which consequently augmented
Jnet (Fig.
5B; P<0.01). These observations were striking because
the ionophore effectively stimulated MeAIB transport (therefore system A
activity) but, as shown before, dramatically reduced net L-alanine fluxes.
Finally, in order to assess the effects of intracellular messengers on the
activity of system ASC, we measured L-alanine fluxes in the presence of a
saturating concentration of MeAIB (20 mmol l1) under
short-circuit conditions and in the presence of sodium. This same procedure
has been used to isolate the ASC transport pathway in different preparations,
including lizard duodenum (Medina et al.,
2001). Interestingly, under these conditions, both intracellular
messengers affected L-alanine transport but in a completely opposite direction
(Figs 4C,
5C). Thus, while db-cAMP
significantly increased Jms and Jnet,
addition of A23187 reduced both Jms and
Jnet. Neither compound affected Jsm,
ruling out a possible change on membrane permeability and indicating that
system ASC activity is stimulated by cAMP and depressed by increased
intracellular free calcium.
Effects of db-cAMP and A23187 on L-alanine- and MeAIB-induced
bioelectrical parameters
To further explore the effects of cAMP and calcium on the activity of the
different L-alanine transport pathways, we designed experiments to ascertain
the electrical responses of isolated duodenum to the addition of transported
substrates before and after addition of the drugs. As can be seen in
Fig. 6A, addition of L-alanine
(4 mmol l1) to the bath readily elevated PD and
Isc, consistent with our previous demonstration of the
activation of Na+- and L-alanine-dependent conductive pathways
(Medina et al., 2001).
Following washout, application of forskolin to the bath induced an increase in
PD and Isc that could be readily reversed by addition of
the chloride channel blocker diphenylamine-2-carboxilic acid (DPC; 1 mmol
l1) but not by addition of
4,4'-diisothiocyanatostilbene-2,2'-disulfonic acid (DIDS; 0.5 mmol
l1; data not shown), suggesting that forskolin had activated
the Cl conductance typically involved in the secretory
response to cAMP-elevating agents (Anderson
and Welsh, 1991
; Anderson et
al., 1992
; Liedtke,
1989
). Under these conditions, subsequent addition of L-alanine
elicited changes on the electrical parameters that were similar in magnitude
to those observed in the control period
(Fig. 6A, inset). These results
indicate that, although forskolin increases unidirectional and net L-alanine
fluxes, the pathway(s) activated by increased cAMP levels do not provide any
additional electrogenicity but, rather, behave in an electroneutral way.
Similar effects were observed using MeAIB instead of L-alanine. These
observations suggest the involvement of an Na+-dependent,
MeAIB-excluding, electroneutral pathway, which may well be ascribed to the
electrical operation of system ASC
(Bussolati et al., 1992
;
Medina et al., 2001
).
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The effects of calcium ionophore on L-alanine-induced electrical responses
suggested that increased intracellular calcium potentiated the response to
L-alanine. Indeed, results illustrated in
Fig. 6B show that exposure of
the tissues to A23187 (0.5 µmol l1) did not significantly
affect either PD or Isc. This observation was interesting
because it ruled out the possible activation of rheogenic calcium-dependent
Cl or K+ conductances in response to the
ionophore (Anderson and Welsh,
1991; Anderson et al.,
1992
; Liedtke,
1989
). However, subsequent addition of L-alanine to A23187-treated
tissues brought about a 23-fold increase in both PD and
Isc, the changes being statistically significant compared
with the first amino acid challenge (Fig.
6B, inset). Identical results were obtained using the system
A-specific substrate MeAIB. Taken together, these results indicate that,
although increased intracellular calcium reduces total L-alanine transport
(see above), it stimulates an electrogenic pathway endowed with properties of
system A.
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Discussion |
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Using electrophysiological and radiotracer techniques, we have recently
demonstrated that active L-alanine transport across the lizard duodenum could
be explained by the simultaneous operation of different carriers
(Díaz et al., 2000;
Medina et al., 2001
). These
distinct transport pathways could be resolved on the basis of their different
Na+ dependence, substrate affinity, kinetic features and electrical
properties. These approaches led us to ascertain that the lizard duodenum
displayed two Na+-dependent pathways, endowed with properties of
system A (MeAIB-transporting electrogenic system) and system ASC
(MeAIB-insensitive L-cysteine- and L-serine-transporting electroneutral
system), which coexist with a unique Na+-independent,
K+-dependent, electrogenic L-alanine transport
pathway.
Under this scenario, we thought it worthwhile to investigate the effects of
increased cAMP and intracellular calcium levels on L-alanine transport
pathways in order to assess whether individual systems were affected equally
by intracellular signal transducers. The data shown in
Fig. 4 suggest that not all
transport pathways exhibited similar responses to db-cAMP; instead, only
system ASC appeared to be stimulated by the nucleotide. The finding that MeAIB
transport was not affected by db-cAMP was striking, since most literature data
on the short-term regulation of neutral amino acid transport by cAMP (or
glucagon) in hepatocytes and adipocytes indicate that system A represents the
main target for cAMP stimulation (Guma et
al., 1993; McGivan et al.,
1981
; Miller and Bhandari,
1986
; Moule et al.,
1987
; Moule and McGivan,
1990
).
Our results also demonstrate that, aside from stimulating net L-alanine
transport, db-cAMP, as well as forskolin and theophylline, increased
transepithelial PD and Isc (see
Table 1). These observations
indicate that the rise in cAMP in duodenal tissues had eventually triggered an
electrogenic process. It has been suggested that rapid stimulation of
L-alanine transport by cAMP in rat hepatocytes was due to membrane
hyperpolarization (Moule et al.,
1987). These authors have suggested that cAMP hyperpolarizes
hepatocytes by stimulation of the Na+/H+-exchange and
Na+/K+-ATPase activities, therefore reducing
intracellular Na+, which would enhance Na+alanine
symport activity in the plasma membrane of hepatocytes
(Moule et al., 1987
). In
addition, numerous studies have demonstrated that these agents stimulate an
electrogenic chloride secretion by activation of apical Cl
channels in the intestinal epithelia (see
Anderson et al., 1992
;
Liedtke, 1989
;
Merlin et al., 1998
). Hence,
evaluation of electrical changes associated with stimulation of L-alanine
transport by increasing cAMP is hampered by the possible induction of an
electrogenic chloride secretion. In order to circumvent this inconvenience, we
designed the sequential protocol shown in
Fig. 6A. Firstly, the effects
of L-alanine on PD and Isc were determined and the
percentage of change was measured (1st addition). Following washout, addition
of forskolin to the bath brought about a considerable increase in PD and
Isc, which was reduced to control levels by DPC, a
well-known inhibitor of cAMP-activated apical Cl channels
(Anderson et al., 1992
;
Becq et al., 1993
). Then, still
in the presence of forskolin and DPC, addition of L-alanine (2nd addition)
rapidly increased PD and Isc, but no appreciable
differences were observed among the two consecutive L-alanine additions,
suggesting that, although L-alanine fluxes were considerably stimulated by
forskolin, the system responsible for this change was not conductive but
rather behaved in an electroneutral manner, ruling out the activation of
system A. In the case of lizard duodenum, this feature defined the electrical
operation of system ASC (Medina et al.,
2001
). Moreover, the magnitude of system ASC activity stimulation
was sufficient to explain the increase in overall L-alanine transport observed
following forskolin. Consistent with our results, studies performed in
skeletal muscle and placental membranes have demonstrated that neither
forskolin nor cholera toxin, both cAMP-elevating factors, affect system A
activity (Guma et al., 1992
;
Pastor-Anglada et al., 1996
).
Likewise, results from trout hepatocytes have revealed that system ASC, but
not system A, was rapidly stimulated by glucagon
(Gallardo et al., 1996
).
Although speculative, our results, together with those from fish hepatocytes,
suggest that rapid regulation of L-alanine transport could have been subjected
to phylogenetic variations.
The experiments aimed at assessing a possible role of calcium on the
regulation of L-alanine transport across the lizard duodenum suggest that
L-alanine transport pathways appear to be oppositely modulated by
intracellular messengers. Indeed, permeabilizing duodenal cells to calcium
using the ionophore A23187 dramatically reduced Jms and
Jnet, without altering Jsm, revealing
that an important fraction of total L-alanine transport was depressed by a
rise in cytosolic calcium. Although the literature on the regulation of amino
acid transport by calcium is scarce, the inhibition of L-alanine and glucose
uptake across brush-border vesicles from rabbit small intestine by
intravesicular calcium loading has been reported by Miyamoto et al.
(1990). Similar findings have
been observed for several Na+-dependent carriers in rat small
intestine vesicles preloaded with a high calcium concentration
(Fondacaro and Madden,
1984
).
Strikingly, in spite of the considerable reduction of duodenal L-alanine
transport by A23187, PD and Isc were augmented by the
ionophore, reflecting the activation of some conductive mechanism (see
Table 2). It is well known that
cytosolic calcium may regulate electrolyte transport in airway and intestinal
epithelia (Brown, 1987;
Liedtke, 1989
) by different
mechanisms, often involving activation of Ca2+-dependent
Cl channels and/or calcium-dependent K+ channels
(Anderson and Welsh, 1991
;
McCabe and Smith, 1985
;
Schultheiss and Diener, 1998
).
However, this does not seem to be the case for the lizard duodenum, as
addition of A23187 to control tissues (in the absence of L-alanine) failed to
produce any significant change in either PD or Isc
(Fig. 6B), therefore ruling out
the possibility of an electrogenic Cl secretion activated by
calcium. The first evidence for the identification of the putative
electrogenic pathway activated by rising intracellular calcium emerged from
the comparison of the effects of L-alanine addition to the lizard duodenum
before and after the ionophore challenge. As can be seen in
Fig. 6B, application of the
calcium ionophore potentiated the responses of PD and Isc
to L-alanine, suggesting that the rise in cytoplasmic calcium had activated an
L-alanine-dependent conductive pathway. Hence, we determined the effects of
calcium ionophore on individual transport pathways using the same dissection
protocol mentioned before for forskolin
(Fig. 5). The results showed
that calcium ionophore dramatically reduced system ASC and augmented system A,
while the Na+-independent pathway remained unaffected.
These observations provided a clue to reconcile the apparent contradiction
that the ionophore reduced total L-alanine transport but increased
L-alanine-induced electrogenicity. Thus, as system ASC represents the most
important contribution to total duodenal L-alanine transport (serving
>70%), its inhibition would account for the dramatic reduction observed on
the overall net L-alanine transfer. On the other hand, as system A catalyses
an electrogenic Na+L-alanine cotransport,
stimulation of system A activity by calcium would lead to enhanced sodium
transfer to the serosal side (most likely involving
Na+/K+-ATPase activity), thereby generating the
concomitant increase in transepithelial PD and Isc. In
agreement with our observations, human T cells demonstrate rapid enhancement
of system A uptake when treated with ionomycin, an effect that was blocked by
extracellular EGTA (Woodlock et al.,
1989). Similar stimulatory actions of calcium on system A activity
have also been observed in endothelial cells
(Escobales et al., 1994
) and
glioblastoma cells (Zafra et al.,
1994
).
In summary, our data provide evidence that L-alanine transport in the lizard intestine is differently modulated by intracellular signal transducers, namely cAMP and calcium. The extent to which each transport pathway is affected by intracellular messengers varies depending on the individual system considered. Thus, while system A appears to be stimulated by elevated intracellular calcium, system ASC activity is clearly reduced. However, system ASC, which represents the largest transport pathway in unstimulated tissues, is clearly stimulated by increased cAMP levels. Although the precise mechanisms of regulation remain elusive, the magnitude of the effects observed here suggests that amino acid transport across the duodenum may be precisely controlled by circulating hormones and paracrine factors reaching the submucosa and the epithelia. Current research is being undertaken to assess the significance of these findings in the context of the physiological responses to endogenous agonists triggering acute changes in the levels of these intracellular messengers.
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Acknowledgments |
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References |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
Anderson, M., Sheppard, D., Berger, H. A. and Welsh, M.
(1992). Chloride channels in the apical membrane of normal and
cystic fibrosis airway and the intestinal epithelia. Am. J.
Physiol. 263,L1
-L4.
Anderson, M. and Welsh, M. (1991). Calcium and cAMP activate different chloride channels in the apical membrane of normal and cystic fibrosis epithelia. Proc. Natl. Acad. Sci. USA 88,6003 -6007.[Abstract]
Becq, F., Hollande, E. and Gola, M. (1993). Phosphorylation-regulated low-conductance Cl channels in a human pancreatic duct cell line. Pflügers Arch. 425, 1-8.[CrossRef][Medline]
Bridges, R. J., Rummel, W. and Simon, B. (1983). Forskolin induced chloride secretion across the isolated mucosa of rat colon descendens. Naunyn-Schmiedeberg's Arch. Pharmacol. 323,355 -360.[Medline]
Brown, D. (1987). Intracellular mediators of peptide action in the intestine and airways: focus on ion transport function. Am. Rev. Respir. Dis. 136,S43 -S48.[Medline]
Bussolati, O., Laris, P. C., Roud, B. M., Dall'Asta, V. and
Gazzola, G. C. (1992). Transport system ASC for neutral amino
acid. J. Biol. Chem.
267,8330
-8335.
Díaz, M. and Cozzi, S. (1991). Fluxplus, a microcomputer program for interactive calculation of "in vitro" solute fluxes, uptakes and accumulation in studies of intestinal transport. Comp. Meth. Prog. Biomed. 34,263 -271.[Medline]
Díaz, M. and Lorenzo, A. (1991). Coexistence of absorptive and secretory NaCl processes in the isolated lizard colon: effects of cyclic AMP. Zool. Sci. 8, 477-484.
Díaz, M., Medina, V., Gómez, T. and Lorenzo,
A. (2000). Membrane mechanisms for electrogenic
Na+-independent L-alanine transport in the lizard duodenal mucosa.
Am. J. Physiol. 279,R925
-R935.
Edmonson, J. W. and Lumeng, L. (1980). Biphasic stimulation of amino acid uptake by glucagon in hepatocytes. Biochem. Biophys. Res. Commun. 96, 61-68.[Medline]
Escobales, N., Martínez, J. and González, O. (1994). Upregulation of Na+-dependent alanine transport in vascular endothelial cells by serum: role of intracellular Ca2+. Microcirculation 1, 49-58.[Medline]
Fondacaro, J. D. and Madden, T. B. (1984). Inhibition of Na+-coupled solute transport by calcium in brush border membrane vesicles. Life Sci. 35,1431 -1438.[CrossRef][Medline]
Gallardo, M., Pesquero, J., Esteve, M., Canals, P. and Sánchez, J. (1996). Regulation of the ASC system and Na+/K+ pump activities in brown trout (Salmon trutta) hepatocytes. J. Exp. Biol. 11,2459 -2465.
Gazzola, G. C., Dall'Asta, V. and Guidotti, G. G.
(1981). Adaptive regulation of amino acid transport in cultured
human fibroblast. J. Biol. Chem.
256,3191
-3198.
Goldstone, A. D., Koening, H., Lu, C. Y. and Trout, J. J. (1983). Beta-adrenergic stimulation evokes a rapid, Ca++-dependent stimulation of endocytosis, hexose and amino acid transport associated with increased Ca++ fluxes in mouse kidney cortex. Biochem. Biophys. Res. Commun. 114,913 -921.[Medline]
Gómez, T., Badía, P., Bolaños, A. and Lorenzo, A. (1986). Transport of galactose and sodium across the lizard duodenum. Comp. Biochem. Physiol. A 85,103 -107.[Medline]
Guma, A., Vinals, F., Testar, X., Palacin, M. and Zorzano, A. (1992). Differential sensitivity of insulin and adaptive regulation induced system A activation to microtubular function in skeletal muscle. Biochem. J. 281,407 -411.[Medline]
Guma, A., Vinals, F., Testar, X., Palacin, M. and Zorzano, A. (1993). Regulation of system A amino acid transport activity by phospholipase C and AMPc-inducing agents in skeletal muscle: modulation of insulin action. Biochem. Biophys. Acta 117,155 -161.
Handlogten, M. E. and Kilberg, M. S. (1984).
Induction and decay of amino acid transport in the liver. Turnover of
transport activity in isolated hepatocytes after stimulation by diabetes or
glucagon. J. Biol. Chem.
259,3519
-3525.
Karl, P., Chang, B. and Fisher, S. E. (1988). Calcium-sensitive uptake of amino acid by human placental slices. Pediatr. Res. 23,9 -13.[Abstract]
Kilberg, M. S. (1986). Amino acid transport in eukaryotic cells and tissues. Fed. Proc. 45,2438 -2454.
Kilberg, M. S., Stevens, B. R. and Novak, D. A. (1993). Recent advances in mammalian amino acid transport. Annu. Rev. Nutr. 13,137 -165.[CrossRef][Medline]
Liedtke, C. M. (1989). Regulation of chloride transport in epithelia. Annu. Rev. Physiol. 51,143 -160.[CrossRef][Medline]
Lorenzo, A., Pérez, A., Badía, P. and Gómez, T. (1989). Influence of theophylline on phenylalanine transport across isolated duodenal mucosa of lizard. Biochem. Int. 18,1051 -1058.
Lorenzo, A., Santana, P., Gómez, T. and Badía, P. (1989). Sodium and chloride transport in the lizard duodenum. Zool. Sci. 6,667 -674.
McCabe, R. D. and Smith, P. L. (1985). Colonic
potassium and chloride secretion: role of cAMP and calcium. Am. J.
Physiol. 248,G103
-G109.
McGivan, J. D. and Pastor-Anglada, M. (1994). Regulatory and molecular aspects of mammalian amino acid transport. Biochem. J. 299,321 -334.[Medline]
McGivan, J. D., Ramsell, J. C. and Lacey, J. H. (1981). Stimulation of alanine transport and metabolism by dibutyryl cyclic AMP in hepatocytes from fed rats. Assessments of transport as a potential rates-limiting step. Biochim. Biophys. Acta 644,295 -304.[Medline]
Medina, V., Lorenzo, A. and Díaz, M.
(2001). Electrogenic Na+-dependent L-alanine transport
in the lizard duodenum. Involvement of systems A and ASC. Am. J.
Physiol. 280,R612
-R622.
Merlin, D., Jiang, L., Strohmeier, G., Nusrat, A., Alper, S.,
Lencer, W. I. and Marada, J. L. (1998). Distinct
Ca2+- and cAMP-dependent anion conductances in the apical membrane
of polarized T84 cells. Am. J. Physiol.
275,C484
-C495.
Miller, R. E. and Bhandari, B. (1986). Hormonal regulation of amino acid uptake by cultured 3T3-L1 adipocytes. Biochem. Int. 12,775 -783.[Medline]
Miyamoto, Y., Kulanthaivel, P., Ganapathy, V., Whitford, G. M. and Leibach, F. H. (1990). Calcium-induced inhibition of taurine transport in brush-border membrane vesicles from rabbit small intestine. Biochim. Biophys. Acta 1030,189 -194.[Medline]
Moule, S. K. and McGivan, J. D. (1990). Epidermal growth factor and cyclic AMP stimulate Na+/H+ exchange in isolated rat hepatocytes. Eur. J. Biochem. 187,677 -682.[Abstract]
Moule, S. K., Bradford, M. N. and McGivan, J. D. (1987). Short term stimulation of Na+-dependent amino acid transport by dibutyryl cyclic AMP in hepatocytes. Characteristics and partial mechanism. Biochem. J. 241,737 -743.[Medline]
Munck, L. K. and Munck, B. G. (1994). Amino acid transport in the small intestine. Physiol. Rev. 43,335 -346.
Pastor-Anglada, M., Felipe, A., Casado, F., Ferrer-Martínez, A. and Gómez-Angelats, M. (1996). Long-term osmotic regulation of amino acid transport system in mammalian cells. Amino Acids 11,135 -151.
Ramamoorthy, S., Leibach, F. H., Mahesh, V. B. and Ganapathy, V. (1992). Modulation of the activity of amino acid transport system L by phorbol esters and calmodulin antagonist in a human placental choriocarcinoma cell line. Biochim. Biophys. Acta 1136,181 -188.[Medline]
Reymann, A., Braun, W. and Woermann, C. (1986). Proabsorptive properties of forskolin: disposition of glycine, leucine and lysine in rat jejunum. Naunyn-Schmiedeberg's Arch. Pharmacol. 334,110 -115.[Medline]
Schultheiss, G. and Diener, M. (1998). K+ and Cl conductances in the distal colon of the rat. Gen. Pharm. 31,337 -342.[CrossRef][Medline]
Shotwell, M. A., Kilberg, M. S. and Oxender, D. L. (1983). Regulation of neutral amino acid transport in mammalian cells. Biochim. Biophys. Acta 737,267 -284.[Medline]
Stevens, B. R. (1992). Vertebrate intestine
apical membrane mechanisms of organic nutrient transport. Am. J.
Physiol. 263,R458
-R463.
Woodlock, T. J., Segel, G. B. and Lichtman, M. A. (1989). Diacylglycerol and calcium induce rapid enhancement of A-system amino acid transport by independent mechanisms in human T-lymphocytes. J. Cell. Physiol. 141, 33-39.[Medline]
Zafra, F., Aragón, C. and Giménez, C. (1994). Characteristics and regulation of proline transport in cultured glioblastoma cells. Biochem. J. 302,675 -680.[Medline]