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Biomedical subjects

M R Williams

Publications and source records attributed to M R Williams.

At least 37 records · Page 2Linked to original sources
PubMed2000

A randomized trial of antithrombin concentrate for treatment of heparin resistance.

BACKGROUND: Heparin resistance is an important clinical problem traditionally treated with additional heparin or fresh frozen plasma. We undertook a randomized clinical trial to determine if treatment with antithrombin (AT) concentrate is effective for treating this condition. METHODS: Patients requiring cardiopulmonary bypass who were considered to be heparin resistant (activated clotting time < 480 seconds after > 450 IU/kg heparin) were randomized to receive either 1000 U AT or additional heparin. RESULTS: AT concentrate was effective in 42 of 44 patients (96%) for immediately obtaining a therapeutic activated clotting time. This compared favorably to 28 of 41 patients (68%) treated with additional heparin (p = 0.001). All patients who failed heparin therapy were successfully treated with AT. The patients receiving AT required less time to obtain an adequate ACT but there was no difference in clinical outcomes among the groups. Study patients had deficient AT activity at baseline (56%+/-25%), which improved in those given AT concentrate (75%+/-31% versus 50%+/-23%, p < 0.0005). CONCLUSIONS: Heparin resistance is frequently associated with AT deficiency. Treating this deficiency with AT concentrate is more effective and faster for obtaining adequate anticoagulation than using additional heparin.

Antithrombins↗
PubMed2000

Recurrent remodeling after ventricular assistance: is long-term myocardial recovery attainable?

BACKGROUND: Long-term left ventricular assist devices (LVAD) have been used both as a bridge to heart transplantation and to recovery of native myocardial function. Despite much evidence for reversal of some of the structural and functional changes present in the failing heart during LVAD support, clinical evidence for sustained myocardial recovery is scant. We describe 2 patients in whom myocardial recovery during LVAD support led to device explanation only to have heart failure recur. This necessitated a second LVAD implantation, a process that we have termed recurrent remodeling. METHODS: The medical records of 2 patients with cardiomyopathy supported with HeartMate LVADs (Thermo Cardiosystems, Inc, Woburn, MA) were retrospectively reviewed. RESULTS: One patient was supported with an LVAD for 2 months, at which time the LVAD was explanted. Progressive deterioration of cardiac function followed, requiring a second LVAD 19 months after LVAD explanation. After 2 months of further LVAD support, a second episode of apparent myocardial recovery was observed during a period of device malfunction. The other patient was supported with an LVAD for 12 months, at which time the LVAD was explanted. The patient experienced progressive hemodynamic deterioration and required a second LVAD 6 months after LVAD explantation. Heart transplantations of both patients were successful. CONCLUSIONS: Our understanding of myocardial recovery in the setting of hemodynamic unloading with LVAD support has not yet progressed to the point where we are able to accurately predict successful long-term LVAD explantation. The evolution of reliable predictors of sustainable myocardial recovery will help to avoid further cases of recurrent remodeling requiring repeat LVAD implantation.

Adult↗
PubMed2000

Arginine vasopressin in the treatment of 50 patients with postcardiotomy vasodilatory shock.

BACKGROUND: The barroreflex-mediated secretion of arginine vasopressin has been found to be defective in a variety of vasodilatory shock states, such as postcardiotomy shock, and administration of the hormone markedly improves vasomotor tone and blood pressure. The high incidence of vasodilatory shock in patients undergoing left ventricular assist device (LVAD) implantation makes this population an ideal model in which to assess the risks and benefits of vasopressin. METHODS: The medical records of the 102 patients receiving LVADs at Columbia-Presbyterian Medical Center from January 1995 to August 1998 were reviewed. Fifty patients were eligible for study based on a history of arginine vasopressin administration in the operating room or intensive care unit within 24 hours of implantation. RESULTS: Despite LVAD implantation and the administration of vasopressors, patients were hypotensive with a mean arterial pressure less than 60 mm Hg. The administration of vasopressin (0.09+/-0.05 U/min) increased mean arterial pressure (58+/-13 to 75+/-14 mm Hg; p<0.001) while reducing norepinephrine administration (11.7+/-13 to 7.9+/-6.0 mcg/min; p = 0.023). There was no significant change in LVAD flow. The incidence of compromised regional perfusion was not different between LVAD patients who received vasopressin as compared to hemodynamically stable LVAD patients who did not receive vasopressin. CONCLUSIONS: We have demonstrated vasopressin at low doses to be a safe and an effective vasopressor in 50 patients with postcardiotomy vasodilatory shock.

Arginine Vasopressin↗
PubMed2000

Six-year experience of caring for forty-four patients with a left ventricular assist device at home: safe, economical, necessary.

OBJECTIVE: With increasing numbers of implantations, left ventricular assist device programs can put a financial strain on a hospital unless an efficient and safe outpatient program is developed. However, the left ventricular assist device is not widely recognized in the medical community as being reliable enough to support a patient at home. We reviewed our experience with these patients at home to assess the safety and the benefits of such a program. METHODS: Our institutional 6-year experience with 90 consecutive recipients of a wearable left ventricular assist device was analyzed. RESULTS: Forty-four (49%) of the 90 patients who received TCI vented-electric left ventricular assist devices (Thermo Cardiosystems, Inc, Woburn, Mass) were discharged, spending a total of 4546 days (12.5 years) at home with an average of 103 +/- 16 days of outpatient support (range 9-436 days). Of these 44 patients, all were successfully bridged to transplantation (42 patients, 96%) or planned explantation (2 patients, 4%). None of the outpatients died. The cumulative events per outpatient month were 0.020 for bleeding, 0.053 for device infection, 0.0068 for thromboembolus, and 0.020 for major malfunctions. Our estimated average cost to bridge a patient to transplantation or explantation once discharged is $13,200 and as an inpatient over the same length of time, including only room and board, is $165,200. Thirty percent of outpatients were able to return to work or school, 33% to sexual activity, and 44% to driving. All outpatients performed activities of daily living. CONCLUSION: Current left ventricular assist device technology provides effective and economical outpatient support and is associated with limited morbidity and a satisfactory quality of life.

Ambulatory Care↗
PubMed1999

Sequence of the genomic RNA of nudaurelia beta virus (Tetraviridae) defines a novel virus genome organization.

The monopartite genome of Nudaurelia beta virus, the type species of the Betatetravirus genus of the family Tetraviridae, consists of a single-stranded positive-sense RNA (ss+RNA) of 6625 nucleotides containing two open reading frames (ORFs). The 5' proximal ORF of 5778 nucleotides encodes a protein of 215 kDa containing three functional domains characteristic of RNA-dependent RNA polymerases of ss+RNA viruses. The 3' proximal ORF of 1836 nucleotides, which encodes the 66-kDa capsid precursor protein, overlaps the replicase gene by more than 99% (1827 nucleotides) and is in the +1 reading frame relative to the replicase reading frame. This capsid precursor is expressed via a 2656-nucleotide subgenomic RNA. The 3' terminus of the genome can be folded into a tRNA-like secondary structure that has a valine anticodon; the tRNA-like structure lacks a pseudoknot in the aminoacyl stem, a feature common to both genera of tetraviruses. Comparison of the sequences of Nudaurelia beta virus and another member of the Tetraviridae, Helicoverpa armigera stunt virus, which is in the genus Omegatetravirus, shows identities of 31.6% for the replicase and 24.5% for the capsid protein. The viruses in the genera Betatetravirus and Omegatetravirus of the Tetraviridae are clearly related but show significant differences in their genome organization. It is concluded that the ancestral virus with a bipartite genome, as found in the genus Omegatetravirus, likely evolved from a virus with an unsegmented genome, as found in the genus Betatetravirus, through evolution of the subgenomic RNA into a separate genomic component, with the accompanying loss of the capsid gene from the longer genomic RNA.

Amino Acid Sequence↗
PubMed1999

Effects of AIF4- and ATP on intracellular calcium dynamics of crypt epithelial cells in mouse small intestine.

Previous digital imaging analysis of intracellular calcium ion ([Ca2+]i) dynamics in the crypt of the small intestine showed little response by most columnar cells to cholinergic and adrenergic agonists. The objective of the present study was to demonstrate whether G-protein activators and other transmitters elicit [Ca2+]i changes in crypt cells. We used digital imaging to analyze spatiotemporal dynamics of [Ca2+]i in Fura-2/AM-loaded isolated crypts of mouse duodenum and ileum. A1F4- increased [Ca2+]i in crypt columnar cells. In many cases, we observed [Ca2+]i oscillations, which were synchronized throughout the entire crypt. The oscillations were blocked by octanol. ATP, but not adenosine, caused a [Ca2+]i increase in middle crypt-regions of the duodenum and upper regions of the ileum, and the [Ca2+]i wave propagated towards the crypt bottom. The ATP-induced [Ca2+]i increase was prevented by pretreatment with thapsigargin or suramin, but not by La3+ or an extracellular Ca(2+)-free environment. Neither dopamine, 5-hydroxytryptamine (5-HT), histamine, vasoactive intestinal peptide, substance P. cholera toxin, nor guanylin had significant effects. The [Ca2+]i dynamics of Paneth cells were independent of the AlF4(-)-induced synchronous oscillations of columnar cells and of the ATP-induced [Ca2+]i wave. In conclusion, crypt columnar cells have [Ca2+]i-dependent intracellular signaling mechanisms that are linked with G proteins, and by which the cells communicate with each other. ATP elicited [Ca2+]i mobilization from columnar cells via P2 receptors, although some regional differences were noted between the duodenum and ileum.

Adenosine Triphosphate↗
PubMed1999

Development of an off bypass mitral valve repair.

BACKGROUND: The Bow Tie Repair (BTR), a single edge-to-edge suture opposing the anterior and posterior leaflets of the mitral valve (MV), has led to satisfactory reduction of mitral regurgitation (MR) with few re-operations and excellent hemodynamic results. The simplicity of the repair lends itself to minimally invasive approaches. A MV grasper has been developed that will coapt both leaflets and fasten the structures with a graduated spiral screw. METHODS: Eleven explanted adult human MVs were mounted in a mock circulatory loop created for simulating a variety of hemodynamic conditions. The MV grasper was used to place a screw in each valve, which was then continuously run for 300,000 to 1,000,000 cycles with a fixed transvalvular pressure gradient. At the completion of these studies, the valves were stressed to a maximal transvalvular gradient for ten minutes. In seven cases, MR was induced and subsequently repaired using the MV screw. In vivo, the MV screw was tested in nine male canines. Through a subcostal incision, the MV grasper entered the left ventricle, approximated the mitral leaflets and deployed the MV screw under direct visualization via an atriotomy. Follow-up transthoracic echocardiograms were done at postoperative week 1, 6, and 12 to identify screw migration, MV regurgitation/stenosis or clot formation. Dogs were sacrificed up to postoperative week 12 to allow gross and histologic assessment. RESULTS: In vitro, no MV screw detached from the valve leaflets or migrated during the durability testing period of 6.8 million cycles, including periods of stress load testing up to 350 mm Hg. The percent regurgitant flow used to assess MR statistically decreased with the placement of the screw from 72 +/- 7% to 34 +/- 17%; p = 0.0025. In vivo, seven dogs whose valves were examined within the first 48 hours revealed leaflet coaptation with an intact MV screw and no evidence of MR. Two dogs, followed for a prolonged period, had serial postoperative echocardiograms demonstrating consistent coaptation, no screw migration, no clot, and no regurgitation or stenosis. In the animal sacrificed at 12 weeks, the MV screw was integrated into the tissue of both leaflets. CONCLUSIONS: The MV screw has provided durable leaflet coaptation and has reduced regurgitation in human MVs. Initial data on the MV screw's biocompatibility and interactions with living valve tissue is promising. Our early success supports further efforts towards the maturation of this prototype into off bypass mitral valve repair technology.

Animals↗
PubMed1998

Acetylcholine-induced calcium signaling along the rat colonic crypt axis.

BACKGROUND & AIMS: Acetylcholine-induced calcium signaling dynamics have been described in cell monolayers derived from colonic mucosa, but not in intact colonic crypts. The aim of this study was to characterize the spatiotemporal characteristics of calcium signaling induced by acetylcholine in isolated intact rat colonic crypts and to identify the muscarinic receptor subtype coupled to this signaling pathway. METHODS: Isolated crypts from the distal colon of male Wistar rats were loaded with the calcium-sensitive dye Fura-2 and imaged with a charge-coupled device video camera. RESULTS: Acetylcholine mobilized intracellular calcium with an EC50 of 3.9 micromol/L. The response was initiated at the base of the crypt and progressed toward the surface. The velocity of propagation was dose dependent. Addition of muscarinic antagonists inhibited the response (pKb values calculated for pirenzepine and 4-DAMP, 6.08 and 8.65, respectively). Microperfusion of acetylcholine initiated a calcium signal throughout the lower half of the crypt. Microinjection of inositol 1,4,5-triphosphate induced a propagation of a calcium signal along the crypt axis. Heptanol inhibited the velocity of acetylcholine-induced wave propagation by 33%. CONCLUSIONS: M3 muscarinic receptors are coupled to the mobilization of calcium from intracellular stores of intact, isolated rat colonic crypts. Intercellular communication potentiates the propagation of the acetylcholine-induced calcium signal along the crypt axis.

Acetylcholine↗
PubMed1998

Platelet transfusions are associated with the development of anti-major histocompatibility complex class I antibodies in patients with left ventricular assist support.

BACKGROUND: Preformed anti-human leukocyte antigen (HLA) antibodies delay heart transplantation in patients with left ventricular assist devices (LVAD) because of difficulty in finding crossmatch-negative donors. These antibodies may also be associated with adverse outcome after transplantation. METHODS: In a retrospective analysis of 40 patients with LVAD at Columbia-Presbyterian Medical Center between 1990 to 1996, age, sex, diagnosis, race, duration of support, transfusions, and infections were studied by univariate and multivariate analysis as predictors for development of either anti-HLA class I (anti-I) or anti-HLA class II (anti-II) immunoglobulin G (IgG) or M (IgM) antibodies. RESULTS: Eighteen (45%) patients had development of anti-I and 20 (50%) had development of anti-II antibodies over the study period. Median time for LVAD support was 142 days (range 35 to 439). Only total number of perioperative platelet transfusions predicted the development of anti-I IgG antibodies (p = .04). No other associations were found for development of anti-I IgM or anti-II antibodies of either IgG or IgM specificity. Patients who had development of anti-I IgG received a mean of 13.9 (SE +/- 2.6) units of platelets compared with a mean of 7.7 (SE +/- 2.3) units in those who did not (p = .01). By Kaplan-Meier analysis, at the median duration of follow-up, 8% of patients receiving < 6 units were predicted to have development of anti-I antibodies compared with 63% receiving > 6 units (p = .002). In the last 7 patients, leukocyte filters were used to decrease the antigenic load during platelet and red blood cell transfusions. Only 1 of 7 (14%) patients had development of anti-HLA antibodies compared with 31 of 33 (94%) in whom filters were not used (p < .005). CONCLUSIONS: These results indicate that platelet transfusion during LVAD implantation is a risk factor associated with development of HLA class I IgG antibodies. Use of leukocyte filters during platelet transfusion may decrease the risk of development of anti-HLA antibodies.

Antibody Formation↗
PubMed1997

Transcriptional regulation of the human alpha1a-adrenergic receptor gene. Characterization Of the 5'-regulatory and promoter region.

We recently cloned cDNAs encoding three subtypes of human alpha1-adrenergic receptors (alpha1ARs), alpha1a, alpha1b, and alpha1d (Schwinn, D. A., Johnston, G. L., Page, S. O., Mosley, M. J., Wilson, K. H., Worman, N. P., Campbell, S., Fidock, M. D., Furness, L. M., Parry-Smith, D. J., Peter, B., and Bailey, D. S. (1995) J. Pharmacol. Exp. Ther. 272, 134-142) and demonstrated predominance of alpha1aARs in many human tissues (Price, D. T., Lefkowitz, R. J., Caron, M. G., Berkowitz, D., and Schwinn, D. A. (1994) Mol. Pharmacol. 45, 171-175). Several lines of evidence indicate that alpha1aARs are important in clinical diseases such as myocardial hypertrophy and benign prostatic hyperplasia. Therefore, we initiated studies to understand mechanisms underlying regulation of alpha1aAR gene transcription. A genomic clone containing 6.2 kb of 5'-untranslated region of the human alpha1aAR gene was recently isolated. Ribonuclease protection and primer extension assays indicate that alpha1aAR gene transcription occurs at multiple initiation sites with the major site located 696 base pairs upstream of the ATG, where a classic initiator sequence is located. Transfection of luciferase reporter constructs containing varying amounts of 5'-untranslated region into human SK-N-MC neuroblastoma cells indicate that a region extending 125 base pairs upstream from the main transcription initiation site contains full alpha1aAR promoter activity. Furthermore, distinct activator and suppressor elements lie 2-3 and 3-5 kilobase pairs upstream, respectively. Although the alpha1aAR promoter contains neither TATA or CAAT elements, gel shift mobility assays targeting three GC boxes immediately upstream of the main transcription initiation site confirm binding of Sp1. Activity of the alpha1aAR promoter is cell-specific, demonstrating highest activity in cells endogenously expressing alpha1aARs. The human alpha1aAR gene also contains several cis regulatory elements, including several insulin and cAMP response elements. Consistent with these observations, we provide the first evidence that treatment of SK-N-MC cells with insulin and cAMP elevating agents leads to an increase in alpha1aAR expression. In conclusion, these data represent the first characterization of the alpha1aAR gene; our findings should facilitate further studies designed to understand mechanisms regulating alpha1AR subtype-specific expression in healthy and diseased human tissue.

Cloning, Molecular↗
PubMed1996

Calcium mobilisation modulates growth of lens cells.

The modulating effect of calcium cell signalling agonists on tissue growth was studied in a rabbit lens cell line (NN1003A). Calcium mobilisation was measured after Fura-2 incorporation and growth assayed either by direct Coulter counting or [3H]-thymidine incorporation. Transient increases in cytoplasmic calcium were elicited by rabbit serum, histamine, ATP and PDGF. Thapsigargin induced a prolonged increase and all of the above agonists failed to elicit a response after thapsigargin. Rabbit serum and PDGF both increased cell growth in a concentration-dependent manner. While histamine and ATP had little effect in serum-free medium, they reduced serum-stimulated growth. Acetylcholine and FGF did not produce a marked rise in cytoplasmic calcium and neither did they modulate growth. Both thapsigargin and caffeine greatly inhibited growth. These findings indicate that, in lens cells, agonists that mobilise calcium, whether by acting through G-protein or tyrosine kinase receptors, also modulate lens cell growth. Agents such as thapsigargin and caffeine that inactivate the same calcium store also inhibit growth.

Acetylcholine↗
PubMed1996

Biventricular assist device as a bridge to transplantation in a pediatric patient.

A 5 1/2-year-old boy with idiopathic cardiomyopathy and rapidly worsening hemodynamic parameters underwent placement of a biventricular assist device as a bridge to transplantation. Direct anastomoses to both the aorta and pulmonary artery with Dacron grafts attached to Carmeda-coated tubing facilitated the support period. Inflow was provided by right atrial appendage and left ventricular apex cannulas. A centrifugal pump provided support for 2 days until a suitable donor was identified. The technique is simple, reproducible, and effective for patients with small body surface areas.

Anastomosis, Surgical↗
PubMed1995

Histamine and ATP mobilize calcium by activation of H1 and P2u receptors in human lens epithelial cells.

1. Cytoplasmic calcium concentration ([Ca2+]1) of single superfused tissue-cultured human lens epithelial cells (HLEC) was monitored using the fluorescent dye fura-2; the resting values were low and stable for several hours ([Ca2+]i = 96 +/- 20 nM; mean +/- S.D., n = 16). 2. Continuous superfusion with either ATP or histamine (0.1-10 microM) produced regular oscillations in [Ca2+]i that could be maintained for a short time in the absence of external calcium. 3. Short (30 s) pulses of histamine (0.1-100 microM) induced a transient rise in [Ca2+]i, the time course of which was insensitive to the removal of external calcium. The rate of rise and the amplitude of the response were very sensitive to agonist concentration, whereas the rate of recovery was relatively constant. 4. The responses to long pulses of histamine (> 100 s) consisted of an initial transient followed by a maintained [Ca2+]i which returned to baseline on removal of external calcium. 5. The kinetics of the responses to short and long pulses of ATP (0.1-100 microM) were very similar to those of histamine and showed a similar sensitivity to the presence or absence of external calcium. 6. The histamine responses were abolished by triprolidine (1 microM), but unaffected by ranitidine (1 microM), indicating that an Hi receptor subtype is activated by histamine. 7. The ATP responses were reversibly inhibited by suramin and the potency sequence for a range of agonists was ATP = UTP = ATP gamma S > ADP = GTP >> AMP = adenosine, indicating that activation of a P2u receptor subtype was responsible for the increase in [Ca2+]i. 8. Both histamine and ATP responses were abolished by thapsigargin (100 nM), confirming that calcium release from intracellular stores was responsible for the initial peak of the response. Application of either agonist during the plateau phase of the thapsigargin response often led to a marked, but reversible, decline in [Ca2+]i, indicating the presence of a further, normally hidden, calcium regulatory factor associated with the presence of the agonist. 9. Maximal concentrations of either histamine or ATP totally emptied the calcium store as a subsequent application of the other agonist (or thapsigargin), in the absence of external calcium, failed to induce a further increase in the calcium signal.

Adenosine Triphosphate↗
PubMed1994

Canine choledochotomy closure with diode laser-activated fibrinogen solder.

BACKGROUND: An alternative to mechanical stapling or hand suturing is needed to permit laparoscopic common bile duct exploration. We evaluated the strength and healing characteristics of canine choledochotomies sealed with a fibrinogen solder and a diode laser. METHODS: After creation of a 0.5 cm longitudinal choledochotomy, the edges were coapted with forceps, and a fibrinogen solder mixed with indocyanine green dye was applied. The solder was sealed in place with an 810 nm diode laser (125 W/cm2). RESULTS: Immediate mean leakage pressure was 264 +/- 7 mm Hg compared with 83 +/- 66 mm Hg in suture controls. This increased to 364 +/- 115 mm Hg at 2 days and was more than 510 mm Hg at 7 days. On histologic examination rapid reabsorption of the solder with no signs of inflammation or stenosis was seen. No episodes of dehiscence or peritonitis occurred. CONCLUSIONS: Laser soldering provides a watertight choledochotomy closure with adequate immediate strength allowing a reliable, technically feasible common bile duct exploration via a laparoscopic approach.

Animals↗
PubMed1993

Laser-assisted fibrinogen bonding of umbilical vein grafts.

Despite success with autologous tissue welding, laser welding of synthetic vascular prostheses has not been possible. The graft material appears inert and fails to allow the collagen breakdown and electrostatic bonding that results in tissue welding. To develop a laser welding system for graft material, we repaired glutaraldehyde-tanned human umbilical cord vein graft incisions using laser-assisted fibrinogen bonding (LAFB) technology. Modified umbilical vein graft was incised transversely (1.2 cm). Incisions were repaired using sutures, laser energy alone, or LAFB. For LAFB, indocyanine green dye was mixed with human fibrinogen and the compound applied with forceps onto the weld site prior to exposure to 808 nm diode laser energy (power density 4.8 W/cm 2). Bursting pressures for sutured repairs (126.6 +/- 23.4 mm Hg) were similar to LAFB anastomoses (111.6 +/- 55.0 mm Hg). No evidence of collateral thermal injury to the graft material was noted. In vivo evaluation of umbilical graft bonding with canine arteries demonstrates that LAFB can reliably reinforce sutured anastomoses. The described system for bonding graft material with laser exposed fibrinogen may allow creation or reinforcement of vascular anastomoses in procedures where use of autologous tissue is not feasible.

Anastomosis, Surgical↗
PubMed1993

Combined modality therapy in the treatment of epidermoid carcinoma of the anus at the Medical University of South Carolina.

Treatment of anal epidermoid carcinoma by radiation and chemotherapy does offer an excellent opportunity to maintain quality of life. No patient in this series suffered long-term complications; and all have maintained rectal, anal, and sexual function. Patients with early stage lesions (AJC I-II) are most amenable to anus conserving therapy for epidermoid cancer of the anus. The success rate is markedly diminished in patients having more advanced disease, with patients possessing AJC stage III and VI disease presenting major problems. The possibility of post operative radiation therapy is markedly compromised by preoperative radiation therapy. Clinical surveillance and early recognition of these lesions by primary physicians can now offer patients with lesser stage cancer of this type excellent opportunity for organ preservation therapy. For those patients presenting with Stage III and VI carcinoma of the anus the aforementioned problems should be addressed and discussed before initiating conservative therapy.

Adult↗
PubMed1992

pH regulation in tissue-cultured bovine lens epithelial cells.

The intracellular pH (pHi) of tissue-cultured bovine lens epithelial cells was measured in small groups of 6 to 10 cells using the trapped fluorescent dye 2',7'-bis-(2-,carboxyethyl)-5 (and 6)carboxyfluorescein (BCECF). When perifused at 35 degrees C with artificial aqueous humour solution (AAH) containing 16 mM HCO3- and 5% CO2, pH 7.25, pH(i) was 7.19 +/- 0.02 (SEM, n = 95). On removing HCO3- and CO2 there was an initial transient alkalinization followed by a fall in pH to a steady value of 6.97 +/- 0.03 (SEM, n = 54). Addition of 0.25 mM 4,4'-diisothiocyanatostilbene-2,2'-disulfonic acid (DIDS) to AAH containing HCO3- and CO2 led to a rapid and pronounced fall in pH. Exposure to Na(+)-free AAH again led to a marked fall in pH(i), but in this case the addition of DIDS did not produce a further fall. Substitution of the impermeant anion gluconate for Cl- in the presence of HCO3- led to a rise in pHi, while substitution in the absence of HCO3- led to a fall in pHi. The above data indicate a significant role for a sodium-dependent Cl(-)-HCO3- exchange mechanism in the regulation of pHi. Addition of 1 mM amiloride to control AAH in both the presence and absence of HCO3- led to a marked fall in pH(i), indicating that a Na+/H+ exchange mechanism also has a significant role in the regulation of pHi.(ABSTRACT TRUNCATED AT 250 WORDS)

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