PubMed Health⌕ Search

Biomedical subjects

Christian Aalkjaer

Publications and source records attributed to Christian Aalkjaer.

At least 19 recordsLinked to original sources
PubMed2006

NBCn1 (slc4a7) mediates the Na+-dependent bicarbonate transport important for regulation of intracellular pH in mouse vascular smooth muscle cells.

The contribution of sodium-dependent bicarbonate transport to intracellular pH (pH(i)) regulation in vascular smooth muscle cells is controversial, partly because the molecular identity of the transporter(s) responsible has not been identified. Here, using the pH-sensitive fluorophore 2',7'-bis-(2-carboxyethyl)-5-(and-6)-carboxyfluorescein (BCECF), we show that smooth muscle cells of intact mouse mesenteric, coronary, and cerebral small arteries all display a sodium- and bicarbonate-dependent pH(i) recovery after an NH4+-prepulse. The sodium-dependent bicarbonate flux was largely 4,4'-diisothiocyanatostilbene-2,2'-disulphonic acid (DIDS) sensitive (56% to 91%) and of a magnitude similar to the amiloride-sensitive flux. Additionally, steady-state pH(i) was lower (0.2 to 0.4 pH units magnitude) in all 3 vascular beds when CO2/bicarbonate was omitted. RT-PCR analyses showed that NBCn1 (slc4a7) is the only Na+-dependent bicarbonate transporter of the slc4 family detectable at the mRNA level in all 3 vascular beds investigated. Whole-mount immunolabeling and immunogold electron microscopy confirmed the presence of NBCn1 protein in the sarcolemma of mouse mesenteric small arterial smooth muscle cells. Intact mouse mesenteric small arteries were electropermeated to facilitate transfection with small interfering RNA targeting NBCn1, which resulted in an approximate 43% decrease in the ratio of NBCn1 to glyceraldehyde-3-phosphate dehydrogenase mRNA. After knock-down, we found a decreased steady-state pH(i) (0.21+/-0.08 pH units) as well as a 68+/-10% decrease in the net Na+-dependent, amiloride-insensitive base influx after acid load. Finally, omission of CO2/bicarbonate resulted in a decreased contractile response to norepinephrine after sustained exposure to the agonist, underlining the importance of CO2/bicarbonate for vascular contractility. We conclude that NBCn1 mediates the Na+-dependent bicarbonate transport important for pH(i) regulation in smooth muscle cells of mouse mesenteric, coronary, and cerebral small arteries.

4,4'-Diisothiocyanostilbene-2,2'-Disulfonic Acid↗
PubMed2006

Analysis of effects of connexin-mimetic peptides in rat mesenteric small arteries.

Synthetic peptides homologous to the extracellular loops of the major vascular connexins represent a novel class of gap junction blockers that have been used to assess the role of direct cellular communication in arteries and veins. However, the specificity of action of such peptides on the coupling between smooth muscle cells (SMCs) has not yet been fully characterized. Isolated third-order rat mesenteric arteries were therefore studied with respect to isometric tension (myography), intracellular Ca2+ concentration ([Ca2+]i) (Ca2+ -sensitive dyes), membrane potential, and input resistance (sharp intracellular glass electrodes). Confocal imaging was used for visualization of [Ca2+]i events in individual SMCs in the arterial wall and membrane currents (patch clamp) measured in individual SMCs isolated from the same arteries. A triple peptide combination (37,43Gap 27 + 40Gap 27 + 43Gap 26) increased intercellular resistance (measured as input resistance) in intact arterial segments without affecting the membrane conductance of individual cells and also interrupted electrical coupling between pairs of rat aortic A7r5 myocytes. In intact arterial segments, the peptides desynchronized [Ca2+]i transients in individual SMCs and abolished vasomotion without suppressing Ca2+ transients in individual cells. They also depolarized SMCs, increased [Ca2+]i, and attenuated acetylcholine-induced, endothelium-dependent smooth muscle hyperpolarization. Experiments with endothelium-denuded arteries suggested that the depolarization produced by the peptides under basal conditions was in part secondary to electrical uncoupling of the endothelium from SMCs with loss of a tonic hyperpolarizing effect of the endothelium. Taken together, the results indicate that connexin-mimetic peptides block electrical signaling in rat mesenteric small arteries without exerting major nonjunctional effects.

Animals↗
PubMed2006

Effect of acidosis on isolated porcine retinal vessels.

PURPOSE: To study the effect of normocapnic (NA) and hypercapnic acidosis (HA) on the tone, the intracellular calcium level ([Ca(2 +)](i)), and the membrane potential of smooth muscle cells in porcine retinal arterioles. METHODS: Twenty-four porcine retinal arterioles were mounted in a wire myograph for isometric recording of the wall tension. The vessels were precontracted with 0.3 microM U46619 and were exposed to NA (pH = 7.0) and HA (pH = 7.0). Intracellular calcium was measured using the fluorophore Fura-2AM (n = 12). In six vessels, 0.1 mM NG-nitroarginine methyl ester (L-NAME) was added to block NO synthesis. The membrane potential of smooth muscles cells was measured in situ with sharp glass electrodes (n = 12). RESULTS: NA and HA induced both a decrease in wall tension from 1.04 +/- 0.06 N/m to 0.65 +/- 0.1 N/m (p < 0.01) (NA) and 0.56 +/- 0.1 N/m (p < 0.01) (HA) and a decrease in [Ca(2 +)](i) as evidenced from the change in the Fura-2 fluorescence emission ratio from 0.66 +/- 0.03 to 0.57 +/- 0.05 (p = 0.005) (NA) and 0.56 +/- 0.05 (p = 0.002) (HA). These results were unaffected by inhibition of NO-synthesis. NA and HA also both induced hyperpolarization of the smooth muscle membrane from -18 +/- 0.7 mV during precontraction to -26 +/- 1.9 mV (p = 0.002) (NA) and -24 +/- 2.6 mV (p = 0.02) (HA). CONCLUSIONS: Acidosis-induced relaxation of the tone in preconstricted isolated porcine retinal arterioles is associated with a decrease in intracellular calcium and a hyperpolarization of the smooth muscle cells. The acidosis-induced relaxation is independent of CO(2) and is not mediated through NO.

Acidosis↗
PubMed2006

Effect of cyclic guanosine-monophosphate on porcine retinal vasomotion.

PURPOSE: Vasomotion refers to periodic oscillations in vascular tone that ensure the intermittent supply of blood to adjacent microvascular units. Previous evidence from vessels outside the eye suggests that cyclic guanosine-monophosphate (cGMP) is involved in the regulation of vasomotion, but it is unknown whether this compound has an effect on vasomotion in retinal vessels. METHODS: Retinal arterioles from porcine eyes were studied in a wire myograph. After initiation of vasomotion, the vessels were stimulated with increasing concentrations of the cGMP agonist 8-Br-cGMP (n = 6), the phosphodiesterase inhibitor zaprinast (n = 6) and the cGMP synthesis inhibitor L-NAME (n = 6). High concentrations of L-NAME blocked vasomotion, and control experiments (n = 20) using 8-Br-cGMP, S-nitroso-N-acetylpenicillamine (SNAP), adenosine and pinacidil were carried out to elucidate whether this effect was related to changes in the general tone of the vessel. Additionally, the relationship between oscillations in vascular tone and intracellular calcium concentration was studied. RESULTS: Induction of cGMP agonistic activity with either 8-Br-cGMP or zaprinast lowered the vasomotion frequency significantly, whereas L-NAME-induced inhibition of cGMP increased this frequency. Neither of the agents affected the amplitude of the oscillations. The control experiments indicated that the effect of cGMP on vasomotion frequency was independent of the accompanying increase in tone. The oscillations in tone during vasomotion were accompanied by similar oscillations in intracellular calcium concentration. CONCLUSION: Cyclic GMP lowers the frequency without affecting the amplitude of vasomotion in isolated porcine retinal arterioles.

Animals↗
PubMed2005

Na+-dependent HCO3- uptake into the rat choroid plexus epithelium is partially DIDS sensitive.

Several studies suggest the involvement of Na+ and HCO3- transport in the formation of cerebrospinal fluid. Two Na+-dependent HCO3- transporters were recently localized to the epithelial cells of the rat choroid plexus (NBCn1 and NCBE), and the mRNA for a third protein was also detected (NBCe2) (Praetorius J, Nejsum LN, and Nielsen S. Am J Physiol Cell Physiol 286: C601-C610, 2004). Our goal was to immunolocalize the NBCe2 to the choroid plexus by immunohistochemistry and immunogold electronmicroscopy and to functionally characterize the bicarbonate transport in the isolated rat choroid plexus by measurements of intracellular pH (pHi) using a dual-excitation wavelength pH-sensitive dye (BCECF). Both antisera derived from COOH-terminal and NH2-terminal NBCe2 peptides localized NBCe2 to the brush-border membrane domain of choroid plexus epithelial cells. Steady-state pHi in choroidal cells increased from 7.03 +/- 0.02 to 7.38 +/- 0.02 (n=41) after addition of CO2/HCO3- into the bath solution. This increase was Na+ dependent and inhibited by the Cl- and HCO3- transport inhibitor DIDS (200 muM). This suggests the presence of Na+-dependent, partially DIDS-sensitive HCO3- uptake. The pHi recovery after acid loading revealed an initial Na+ and HCO3- -dependent net base flux of 0.828 +/- 0.116 mM/s (n = 8). The initial flux in the presence of CO2/HCO3- was unaffected by DIDS. Our data support the existence of both DIDS-sensitive and -insensitive Na+- and HCO3- -dependent base loader uptake into the rat choroid plexus epithelial cells. This is consistent with the localization of the three base transporters NBCn1, Na+-driven Cl- bicarbonate exchanger, and NBCe2 in this tissue.

4,4'-Diisothiocyanostilbene-2,2'-Disulfonic Acid↗
PubMed2005

Distribution of cGMP-dependent and cGMP-independent Ca(2+)-activated Cl(-) conductances in smooth muscle cells from different vascular beds and colon.

In the present patch-clamp study we have, for the first time, shown the tissue distribution of a recently characterized cGMP-dependent Ca(2+)-activated Cl(-) conductance [18] in smooth muscle cells freshly isolated from different regions: aorta, pulmonary artery, tail artery, femoral artery, femoral vein, middle cerebral artery, renal artery, portal vein, superior mesenteric artery, mesenteric small artery and colon. The cGMP-dependent Cl(-) conductance has properties distinct from those of the 'classical' Ca(2+)-activated Cl(-) conductances; their different sensitivities to niflumic acid and zinc were here utilized to distinguish them. They were found to be co-expressed in different patterns in smooth muscle cells of different origins. The cGMP-dependent conductance was greater in myocytes from cerebral artery and femoral vein and was greater in the renal artery, aorta, mesenteric small artery, femoral artery and the superior mesenteric artery. The presence of the cGMP-dependent Ca(2+)-activated Cl(-) current in smooth muscle cells isolated from the colon demonstrates that this conductance is not limited to the vasculature. The 'classical' Ca(2+)-activated Cl(-) conductance was strongly expressed in smooth muscle cells from the portal vein and the tail artery, and noticeably higher in the pulmonary artery.

Animals↗
PubMed2005

Effects of cGMP on coordination of vascular smooth muscle cells of rat mesenteric small arteries.

OBJECTIVE: We tested the hypothesis that cGMP can induce a state of only partial coordination of vascular smooth muscle cells (VSMC). METHODS: This was done by studying the concentration-dependent effect of 8Br-cGMP on isometric and isobaric force development of noradrenaline-activated segments of rat mesenteric small arteries in which the endothelium was removed. We further measured the concentration-dependent effect of 8Br-cGMP on VSMC membrane potential, spatially resolved [Ca(2+)](i) and VSMC membrane conductance. RESULTS: With 300 microM 8Br-cGMP, coordinated [Ca(2+)](i) activity and vasomotion were seen as previously reported. At 10-30 microM 8Br-cGMP, beating isometric tension oscillations were seen. Isobaric recordings revealed oscillations with different frequencies in different parts of the arteries. At these (10-30 microM) 8Br-cGMP concentrations, membrane potential oscillations did not always concur with isometric tension oscillations, and [Ca(2+)](i) oscillations were only synchronized locally within groups of cells. 8Br-cGMP concentration-dependently decreased the frequency of vasomotion and, in unsynchronized hyperpolarized VSMC, the frequency of [Ca(2+)](i) waves. CONCLUSION: Our results demonstrated that cGMP can cause a partial coordination of the VSMC in the vascular wall (and at high concentrations near complete coordination). Furthermore, the cGMP concentration-dependent decrease of Ca(2+) wave frequency and of vasomotion frequency suggests that cGMP modifies oscillatory Ca(2+) release from the sarcoplasmic reticulum and supports the suggestion that this oscillatory release paces vasomotion.

Animals↗
PubMed2005

Phase resetting of arterial vasomotion by burst stimulation of perivascular nerves.

Arteries display cyclic diameter variations, vasomotion. In vivo, these rhythmic contractions are modulated by the influence of sympathetic nerves. In this study, we investigated the effect of burst stimulation of intramural nerves in vitro on the vasomotion of rat mesenteric small arteries. Vessels were mounted for isometric force measurement. After initiation of vasomotion with noradrenaline (0.5-2 microM), periarterial sympathetic nerves were stimulated electrically (10 impulses at 20 Hz) at approximately half-minute intervals. With a delay of 2-3 s, a neurogenic burst caused a brief contraction of the vascular smooth muscle and altered the period of the current vasomotion cycle. The effect on amplitude decayed rapidly and was practically not apparent in the next vasomotion cycle after the burst. With respect to period, stimulation at increasing intervals from the trough in force of vasomotion caused gradual prolongation of the cycle until a critical interval was reached, after which cycle duration was reduced instead. Since subsequent cycles were not affected, a change in phase remained. When two segments of oscillating arteries were mounted in a two-vessel myograph, simultaneously applied bursts of impulses synchronized their oscillation. The data suggest that changes in neural activity are able to make different vessels oscillate in phase, thereby coordinating vasomotion in different parts of the vascular tree, possibly explaining the synchronicity of vasomotion in different vascular beds that can be observed in vivo.

Animals↗
PubMed2005

Vasomotion: cellular background for the oscillator and for the synchronization of smooth muscle cells.

1. Vasomotion is the oscillation of vascular tone with frequencies in the range from 1 to 20 min(-1) seen in most vascular beds. The oscillation originates in the vessel wall and is seen both in vivo and in vitro. 2. Recently, our ideas on the cellular mechanisms responsible for vasomotion have improved. Three different types of cellular oscillations have been suggested. One model has suggested that oscillatory release of Ca2+ from intracellular stores is important (the oscillation is based on a cytosolic oscillator). A second proposed mechanism is an oscillation originating in the sarcolemma (a membrane oscillator). A third mechanism is based on an oscillation of glycolysis (metabolic oscillator). For the two latter mechanisms, only limited experimental evidence is available. 3. To understand vasomotion, it is important to understand how the cells synchronize. For the cytosolic oscillators synchronization may occur via activation of Ca2+-sensitive ion channels by oscillatory Ca2+ release. The ensuing membrane potential oscillation feeds back on the intracellular Ca2+ stores and causes synchronization of the Ca2+ release. While membrane oscillators in adjacent smooth muscle cells could be synchronized through the same mechanism that sets up the oscillation in the individual cells, a mechanism to synchronize the metabolic-based oscillators has not been suggested. 4. The interpretation of the experimental observations is supported by theoretical modelling of smooth muscle cells behaviour, and the new insight into the mechanisms of vasomotion has the potential to provide tools to investigate the physiological role of vasomotion.

Animals↗
PubMed2004

KATP-channel-induced vasodilation is modulated by the Na,K-pump activity in rabbit coronary small arteries.

The purpose of the study was to evaluate the importance of the Na,K-pump in relaxations induced by K(ATP)-channel openers in rabbit coronary small arteries. Arterial segments were mounted in myographs for recording of isometric tension. Whole-cell patch clamp was used to assess K(ATP)-channel currents in isolated smooth muscle cells from the arteries. In arteries preconstricted with the thromboxane A(2) analogue U46619 pinacidil and cromakalim induced concentration-dependent relaxations. In arteries preconstricted with potassium (124 mM) only high concentrations of pinacidil had a small relaxant effect. In arteries preconstricted with U46619 pinacidil-induced relaxations were unaffected by pretreatment with N(omega)-nitro-L-arginine (L-NNA) and only slightly reduced after mechanical removal of the endothelium. Pinacidil induced relaxations were not significantly affected by 1 microM glibenclamide. However, the relaxations were partly inhibited in potassium-free media and by 1 microM ouabain. In contrast, the concentration-dependent relaxation to cromakalim was partly blocked by 1 microM glibenclamide and partly by 1 microM ouabain and when both drugs were present the inhibition increased. Ouabain (1 microM) and glibenclamide (1 microM) each partly inhibited an ATP-sensitive current induced by pinacidil and cromakalim. In the presence of both inhibitors a greater inhibition was seen. When the solution in the patch pipette was sodium-free the current was reduced and ouabain had no effect. The study suggests that the relaxation to cromakalim and most likely pinacidil is mediated through opening of K(ATP) channels. Inhibition of the Na,K-pump, however, may change the local environment for the K(ATP) channels (i.e. increases the ATP/ADPratio and/or decreases the transmembrane potassium gradient), which partly prevents the activation of the K(ATP)-channel current.

ATP-Binding Cassette Transporters↗
PubMed2004

Functional up-regulation of basolateral Na+-dependent HCO3- transporter NBCn1 in medullary thick ascending limb of K+-depleted rats.

The electroneutral, Na(+)-coupled HCO(3)(-) co-transporter (NBCn1) is expressed in the basolateral membrane of rat medullary thick ascending limb (mTAL) segments and is up-regulated strongly during metabolic acidosis. We have suggested previously that NBCn1-mediated HCO(3)(-) uptake across the basolateral cell membrane is an important buffering mechanism during the transcellular transport of NH(4)(+) in the mTAL. To investigate this further we treated rats with a low-K(+) diet for 4 days, a manoeuvre known to increase proximal tubular ammoniagenesis and delivery of luminal NH(4)(+) to the mTAL. Hypokalaemia strongly increased immunolabelling of basolateral NBCn1 in the mTAL of the inner stripe of the outer medulla. Immunoblotting revealed an eightfold up-regulation of NBCn1 protein in K(+)-depleted rats. Subsequently we used in vitro perfusion of isolated mTAL and BCECF imaging to investigate Na(+)-coupled HCO(3)(-) influx in normal and K(+)-depleted rats. Hypokalaemia induced a threefold up-regulation of Na(+)-coupled HCO(3)(-) influx (1.64+/-0.28 vs. 0.47+/-0.05 pH units/minute, n=8). The Na(+)-dependent alkalinization was also significantly larger in K(+)-depleted rats (0.38+/-0.04 vs. 0.23+/-0.03 pH units). These data indicate that K(+)-depleted rats respond with a strong up-regulation of NBCn1 protein and function, probably to cope with the higher tubular load of NH(4)(+), and strengthen our previous suggestion that NBCn1 is an important player in the excretion of NH(4)(+). They also indicate that it is the delivery of luminal NH(4)(+) rather than systemic changes of pH that determine the expression of NBCn1.

Animals↗
PubMed2004

Junctional and nonjunctional effects of heptanol and glycyrrhetinic acid derivates in rat mesenteric small arteries.

1 Heptanol, 18alpha-glycyrrhetinic acid (18alphaGA) and 18beta-glycyrrhetinic acid (18betaGA) are known blockers of gap junctions, and are often used in vascular studies. However, actions unrelated to gap junction block have been repeatedly suggested in the literature for these compounds. We report here the findings from a comprehensive study of these compounds in the arterial wall. 2 Rat isolated mesenteric small arteries were studied with respect to isometric tension (myography), [Ca2+]i (Ca(2+)-sensitive dyes), membrane potential and--as a measure of intercellular coupling--input resistance (sharp intracellular glass electrodes). Also, membrane currents (patch-clamp) were measured in isolated smooth muscle cells (SMCs). Confocal imaging was used for visualisation of [Ca2+]i events in single SMCs in the arterial wall. 3 Heptanol (150 microm) activated potassium currents, hyperpolarised the membrane, inhibited the Ca2+ current, and reduced [Ca2+]i and tension, but had little effect on input resistance. Only at concentrations above 200 microm did heptanol elevate input resistance, desynchronise SMCs and abolish vasomotion. 4 18betaGA (30 microm) not only increased input resistance and desynchronised SMCs but also had nonjunctional effects on membrane currents. 18alphaGA (100 microm) had no significant effects on tension, [Ca2+]i, total membrane current and synchronisation in vascular smooth muscle. 5 We conclude that in mesenteric small arteries, heptanol and 18betaGA have important nonjunctional effects at concentrations where they have little or no effect on intercellular communication. Thus, the effects of heptanol and 18betaGA on vascular function cannot be interpreted as being caused only by effects on gap junctions. 18alphaGA apparently does not block communication between SMCs in these arteries, although an effect on myoendothelial gap junctions cannot be excluded.

Animals↗
PubMed2004

NBCn1 is a basolateral Na+-HCO3- cotransporter in rat kidney inner medullary collecting ducts.

Primary cultures of rat inner medullary collecting duct (IMCD) cells Na(+) dependently import HCO(3)(-) across the basolateral membrane through an undefined transport protein. We used RT-PCR, immunoblotting, and immunohistochemistry to identify candidate proteins for this basolateral Na(+)-HCO(3)(-) cotransport. The mRNA encoding the electroneutral Na(+)-HCO(3)(-) cotransporter NBCn1 was detected as the only Na(+)-HCO(3)(-) cotransporter in the rat inner medulla (IM) among the five characterized Na(+)-dependent HCO(3)(-) transporters. The mRNA of a yet uncharacterized transporter-like protein, BTR1, was also present in the IM, but its expression in microdissected tubules seemed restricted to the thin limbs of Henle's loop. Immunoblotting confirmed the presence of NBCn1 as an approximately 180-kDa protein of the rat IM. Anti-NBCn1 immunolabeling was confined to the basolateral plasma membrane domain of IMCD cells in the papillary two-thirds of the IM. Consistent with the presence of NBCn1, IMCD cells possessed stilbene-insensitive, Na(+)- and HCO(3)(-)-dependent pH recovery after acidification, as assessed by fluorescence microscopy using a pH-sensitive intracellular dye. In furosemide-induced alkalotic rats, NBCn1 protein abundance was decreased in both the IM and inner stripe of outer medulla (ISOM) as determined by immunoblotting and immunohistochemistry. In contrast, NBCn1 abundance in the IM and ISOM was unchanged in NaHCO(3)-loaded animals, and the NBCn1 abundance increased only in the ISOM after NH(4)Cl loading. In conclusion, NBCn1 is a basolateral Na(+)-HCO(3)(-) cotransporter of IMCD cells and is differentially regulated in IMCD and medullary thick ascending limb.

4,4'-Diisothiocyanostilbene-2,2'-Disulfonic Acid↗
PubMed2004

A cyclic GMP-dependent calcium-activated chloride current in smooth-muscle cells from rat mesenteric resistance arteries.

We have previously demonstrated the presence of a cyclic GMP (cGMP)-dependent calcium-activated inward current in vascular smooth-muscle cells, and suggested this to be of importance in synchronizing smooth-muscle contraction. Here we demonstrate the characteristics of this current. Using conventional patch-clamp technique, whole-cell currents were evoked in freshly isolated smooth-muscle cells from rat mesenteric resistance arteries by elevation of intracellular calcium with either 10 mM caffeine, 1 microM BAY K8644, 0.4 microM ionomycin, or by high calcium concentration (900 nM) in the pipette solution. The current was found to be a calcium-activated chloride current with an absolute requirement for cyclic GMP (EC50 6.4 microM). The current could be activated by the constitutively active subunit of PKG. Current activation was blocked by the protein kinase G antagonist Rp-8-Br-PET-cGMP or with a peptide inhibitor of PKG, or with the nonhydrolysable ATP analogue AMP-PNP. Under biionic conditions, the anion permeability sequence of the channel was SCN- > Br- > I- > Cl- > acetate > F- >> aspartate, but the conductance sequence was I- > Br- > Cl- > acetate > F- > aspartate = SCN-. The current had no voltage or time dependence. It was inhibited by nickel and zinc ions in the micromolar range, but was unaffected by cobalt and had a low sensitivity to inhibition by the chloride channel blockers niflumic acid, DIDS, and IAA-94. The properties of this current in mesenteric artery smooth-muscle cells differed from those of the calcium-activated chloride current in pulmonary myocytes, which was cGMP-independent, exhibited a high sensitivity to inhibition by niflumic acid, was unaffected by zinc ions, and showed outward current rectification as has previously been reported for this current. Under conditions of high calcium in the patch-pipette solution, a current similar to the latter could be identified also in the mesenteric artery smooth-muscle cells. We conclude that smooth-muscle cells from rat mesenteric resistance arteries have a novel cGMP-dependent calcium-activated chloride current, which is activated by intracellular calcium release and which has characteristics distinct from other calcium-activated chloride currents.

Animals↗
PubMed2003

Basolateral Na+-dependent HCO3- transporter NBCn1-mediated HCO3- influx in rat medullary thick ascending limb.

The electroneutral Na(+)-dependent HCO3- transporter NBCn1 is strongly expressed in the basolateral membrane of rat medullary thick ascending limb cells (mTAL) and is up-regulated during NH4(+)-induced metabolic acidosis. Here we used in vitro perfusion and BCECF video-imaging of mTAL tubules to investigate functional localization and regulation of Na(+)-dependent HCO3- influx during NH4(+)-induced metabolic acidosis. Tubule acidification was induced by removing luminal Na+ (DeltapHi: 0.88 +/- 0.11 pH units, n = 10). Subsequently the basolateral perfusion solution was changed to CO2/HCO3- buffer with and without Na+. Basolateral Na(+)-H+ exchange function was inhibited with amiloride. Na(+)-dependent HCO3- influx was determined by calculating initial base flux of Na(+)-mediated re-alkalinization. In untreated animals base flux was 8.4 +/- 0.9 pmol min(-1) mm(-1). A 2.4-fold increase of base flux to 21.8 +/- 3.2 pmol min(-1) mm(-1) was measured in NH4(+)-treated animals (11 days, n = 11). Na(+)-dependent re-alkalinization was significantly larger when compared to control animals (0.38 +/- 0.03 versus 0.22 +/- 0.02 pH units, n = 10). In addition, Na(+)-dependent HCO3- influx was of similar magnitude in chloride-free medium and also up-regulated after NH4+ loading. Na(+)-dependent HCO3- influx was not inhibited by 400 microm DIDS. A strong up-regulation of NBCn1 staining was confirmed in immunolabelling experiments. RT-PCR analysis revealed no evidence for the Na(+)-dependent HCO3- transporter NBC4 or the two Na(+)-dependent CI-/HCO3- exchangers NCBE and NDCBE. These data strongly indicate that rat mTAL tubules functionally express basolateral DIDS-insensitive NBCn1. Function and protein are strongly up-regulated during NH4(+)-induced metabolic acidosis. We suggest that NBCn1-mediated basolateral HCO3- influx is important for basolateral NH3 exit and thus NH4+ excretion by means of setting pHi to a more alkaline value.

Animals↗
PubMed2003

Characterization of vasomotion in porcine retinal arterioles.

PURPOSE: To characterize vasomotion in porcine retinal arterioles in vitro using isobaric (pressure myograph) and isometric (wire myograph) methods. METHODS: Pressure myograph: 208 small porcine retinal arterioles (outer diameter 68 +/- 4 microm) were studied under isobaric conditions in a double-barrelled pipette system. Diameter changes of the arterioles were registered by video recordings. Wire myograph: 60 large porcine retinal arterioles (inner diameter 147 +/- 1.6 microm) were studied under isometric conditions in a small vessel myograph for force measurements. RESULTS: The rates of success in initiating vasomotion were 7.2% using the pressure myograph and 43% using the wire myograph (p < 0.001). The small vessels studied under isobaric conditions oscillated with a frequency of 0.014 Hz and the episodes lasted 6.0 +/- 1.0 min, whereas the large vessels under isometric conditions oscillated with a significantly faster frequency of 0.043 Hz and lasted 32.1 +/- 4.9 min (p = 0.026). CONCLUSION: Retinal vasomotion can be studied in vitro using both pressure myograph and wire myograph techniques. The wire myograph is superior to the pressure myograph in initiating and maintaining vasomotion in vitro.

Animals↗
PubMed2003

Myogenic response in isolated porcine retinal arterioles.

PURPOSE: To study the myogenic response in large and small porcine retinal arterioles and the effects of blockade of L-type voltage gated calcium channels on the myogenic response. METHODS: Eleven large (outer diameter 137.7 +/- 4.5 micro m) and fourteen small (77.3 +/- 3.7 micro m) isolated porcine retinal arterioles were studied for myogenic response while mounted between two double-barrelled pipette systems. The intraluminal pressure was varied from 0 to 140 cm H( 2)O (0-103 mmHg) and the diameter changes of the arterioles were registered. The pressure-diameter relation was determined for arterioles in calcium-containing and calcium free solutions and during inhibition of the L-type voltage gated calcium channels with nifedipine. RESULTS: Both large and small retinal arterioles showed a decrease in diameter as the intraluminal pressure was increased, but no significant difference was found between large and small arterioles (p = 0.39). Blockade of voltage gated calcium channels significantly abolished the myogenic response (p = 0.010 and p = 0.005, large and small arterioles respectively). CONCLUSIONS: The results indicate that myogenic responses are present throughout the retinal arteriolar system and that L-type voltage gated calcium channels are involved in transforming the stretch of retinal vascular smooth muscle cells induced by an increase in intraluminal pressure into a contraction.

Animals↗