Isotonic NaCl Solution Research Articles

509. Non-Viral Gene Delivery in the Murine Nasal Epithelium by Hypotonic Shock

Cystic fibrosis is the most frequent genetic disease among Caucasians. It is caused by mutations in the Cystic Fibrosis Transmembrane conductance Regulation (CFTR) gene. Several viral and non-viral methods have led to insufficient gene transfers in the nasal epithelium, in animal models and in clinic. Moreover, viral vectors elicit a strong immune response following their first administration in the nose, rending impossible any further delivery. In comparison with an isotonic solution of NaCl, the nasal perfusion of naked DNA in deionized water results in a two-order of magnitude higher luciferase activity (2x105 RLU/mg prot.). Furthermore, the perfusion of deionized water 20 minutes prior the perfusion of naked DNA in deionized water leads to a one-order of magnitude reduction of luciferase activity. We hypothesize that deionized water induces a hypotonic shock of nasal epithelial cells. The first step of hypotonic shock consists of an entry of water triggering an intensive exocytosis, allowing the expansion of plasma membrane. This latter might therefore resist to cell swelling without breaking. The second step is called Regulatory Volume Decrease (RVD) and basically consists in retrieving the excess plasma membrane by intensive endocytosis. It is believed that plasmid DNA, bound on the apical surface of epithelial cells, would be taken up during the RVD. While nasal epithelial cells are still recovering from a first hypotonic shock, a second hypotonic shock would not induce an uptake of plasmid DNA as intensive as in the course of a single hypotonic treatment. Hypotonic shock-mediated local gene delivery might be applicable in tissues other than nasal epithelium. The inhibition of DNases by Aurintricarboxylic acid (20 μg per mouse) led to a 3.4-fold improvement of luciferase activity (6.7x105 RLU/mg prot.), that is the highest achieved with a non-viral gene delivery system in the nasal epithelium, so far.

Cobalt-Protoporphyrin treatment renders islets tolerant to interleukin-1 beta suppression

This study examined whether treating donor mice with a single dose of cobalt protoporphyrin (CoPP) induced heme oxygenase-1 (HO-1) and protected islet cells from interleukin-1 beta (IL-1β) suppression. Islets were isolated from mice receiving a single dose of either CoPP (20 mg/kg of body weight, CoPP islets) or isotonic NaCl solution vehicle (control islets), 24 hours before isolation. Glucose-stimulated insulin secretion (GSIS) and insulin content (IC) of the islets were determined following incubation in the presence versus absence of murine IL-1β for 21 or 65 hours. The HO-1 protein level of CoPP-induced islets, as determined by an enzyme immunoassay, was significantly higher than that of control islets at 12 hours ( P < .01) and 30 hours ( P < .05), and returned to basal levels at 56 hours ( P = NS). Following a 21-hour incubation with IL-1β, CoPP islets secreted significantly more insulin upon glucose stimulation and preserved significantly more IC than control islets. After 65-hour incubation with IL-1β, CoPP islets secreted significantly less insulin upon glucose stimulation than control islets and preserved significantly less IC compared to islets incubated without IL-1β. In conclusion, treatment with cobalt-protoporphyrin to induce heme oxygenase-1 protects islets against the suppressive effects of IL-1β.

Spectral-Luminescent Properties of Cationic Indotricarbocyanine Dyes in Cancerous Cells and Solutions

The spectral-luminescent properties of symmetrical indotricarbocyanine dyes with the same cation and different anions (Br−, BF4 −) in cancerous HeLa cells, an isotonic solution of NaCl, and organic solvents have been investigated. It is shown that when these dyes are present in cells in nontoxic concentrations, they are in a monomeric state, they are localized in the region with a low dielectric permeability, bonded to biological macromolecules, and do not make contact with an aqueous medium. It has been established for the first time that the molecules of the polymeric dyes are present in cells predominantly in the form of contact ion pairs, and in a dye with the Br− anion the presence of free cations or solvately separated ionic pairs is revealed. It is shown that the shift of the ion equilibrium toward contact ion pairs for the dyes in the cells (as compared to the low-polarity organic solvents) can be due to the fairly high concentration of salts in the biosystem.

Iron and magnesium exchange via the low affinity iron transporter in rabbit erythroid cells—exchange rates and the action of valinomycin, diethylstilbestrol and protein kinase inhibitors

Evidence was presented previously that rabbit erythroid cells possess a low-affinity Fe2+ transport system which operates via the Na+/Mg2+ antiport [Biochim. Biophys. Acta 1282 (1996) 163]. This was investigated further by measurements of Mg2+ efflux as well as Fe2+ uptake by the cells and by examining the inhibitory effects of valinomycin, diethylstilbestrol (DES) and protein kinase inhibitors. Mg2+ efflux and Fe2+ uptake were measured using rabbit reticulocytes and mature erythrocytes incubated in isotonic KCl or NaCl solutions. Both processes were slower in mature cells than reticulocytes. Mg2+ efflux into KCl solution was much lower than into NaCl solution but was stimulated by addition of Fe2+ to the solution. The rate of Fe2+-stimulated Mg2+ efflux closely followed that of Fe2+ uptake in a one-to-one molar ratio. Valinomycin, DES and the protein kinase inhibitors all inhibited Fe2+ uptake from KCl solution. Valinomycin also inhibited Fe2+-stimulated Mg2+ efflux into KCl solution but markedly stimulated the efflux into NaCl. Maximal inhibition of Fe2+ uptake from KCl solution required the presence of K+, Rb+ or Cs+ ions with which valinomycin forms strong complexes. The results could not be explained on the basis of changes in cell membrane potential or cell volume. By contrast, the increase in Mg2+ efflux into NaCl solution produced by valinomycin was accompanied by cell shrinkage and production of a more negative membrane potential, either of which may be responsible for the effect. The inhibition produced by the protein kinase inhibitors indicate that phosphorylation of the transporter or an associated protein by protein tyrosine kinase is probably required to activate the transporter.

Stimulation of potassium cycling in mitochondria by long-chain fatty acids

Nonesterified long-chain fatty acids (myristic, palmitic, oleic and arachidonic), added at low amounts (around 20 nmol/mg protein) to rat liver mitochondria, energized by respiratory substrates and suspended in isotonic solutions of KCl, NaCl, RbCl or CsCl, adjusted to pH 8.0, induce a large-scale swelling followed by a spontaneous contraction. Such swelling does not occur in alkaline solutions of choline chloride or potassium gluconate or sucrose. These changes in the matrix volume reflect a net uptake, followed by net extrusion, of KCl (or another alkali metal chloride) and are characterized by the following features: (1) Lowering of medium pH from 8.0 to 7.2 results in a disappearance of the swelling-contraction reaction. (2) The contraction phase disappears when the respiration is blocked by antimycin A. (3) Quinine, an inhibitor of the K +/H + antiporter, does not affect swelling but suppresses the contraction phase. (4) The swelling phase is accompanied by a decrease of the transmembrane potential and an increase of respiration, whereas the contraction is followed by an increase of the membrane potential and a decrease of oxygen uptake. (5) Nigericin, a catalyst of the K +/H + exchange, prevents or partly reverses the swelling and partly restores the depressed membrane potential. These results indicate that long-chain fatty acids activate in liver mitochondria suspended in alkaline saline media the uniporter of monovalent alkali metal cations, the K +/H + antiporter and the inner membrane anion channel. These effects are presumably related to depletion of mitochondrial Mg 2+, as reported previously [Arch. Biochem. Biophys. 403 (2002) 16], and are responsible for the energy-dissipating K + cycling. The uniporter and the K +/H + antiporter are in different ways activated by membrane stretching and/or unfolding, resulting in swelling followed by contraction.

Melatonin reduces lipid peroxidation and nitric oxide during irradiation-induced oxidative injury in the rat liver.

Radiation therapy is a popular and useful tool in the treatment of cancer. Melatonin participates in the regulation of a number of important physiological and pathological processes. Melatonin, a powerful endogenous antioxidant, plays a role in the reduction of oxidative damage. Thirty adult rats were divided into five equal groups. On the day of the experiment, groups I and II were injected with 5 or 10 mg/kg melatonin, respectively, while group III received isotonic NaCl solution. Thirty minutes later, groups I, II and III were exposed to 6.0 Gy whole body ionizing radiation in a single fraction. Group IV was injected with 5 mg/kg melatonin but was not irradiated. The final group was reserved as sham treated. Liver malondialdehyde (MDA) and nitric oxide (NO*) levels were measured in all groups. Whole body irradiation caused a significant increase in liver MDA and NO* levels. Hepatic MDA and NO* levels in irradiated rats that were pretreated with melatonin (5 or 10 mg/kg) were significantly decreased. Malondialdehyde and NO* levels were reduced in a dose-related manner by melatonin. The data show that melatonin reduces liver damage inflicted by irradiation when given prior to the exposure to ionizing radiation. The radioprotective effect of melatonin is likely achieved by its ability to function as a scavenger for free radicals generated by ionizing radiation.

Effects of glucagon-like peptide-1 (7–36) amide on neurohypophysial hormone secretion induced by acute hyperosmotic challenge

This study was designed to investigate possible effects of glucagon-like peptide-1 (7–36) amide on the vasopressin and oxytocin release induced by acute peripheral or central osmotic stimulation. In the first series of experiments, rats were injected intraperitoneally with the isotonic (0.15 M) or hypertonic (1.5 M) NaCl solution and then, intracerebroventricularly, with either 1 μg glucagon-like peptide-1 (7–36) amide dissolved in 5 μl of isotonic saline or with the vehicle only. In the second study, 1 μg glucagon-like peptide-1 (7–36) amide, dissolved in isotonic or hypertonic (0.6 M) saline, was injected into the cerebroventricular system. Control rats were treated with isotonic or hypertonic saline only. All the animals were decapitated 10 min after the intracerebroventricular injection. Glucagon-like peptide-1 (7–36) amide enhanced significantly the basal secretion of vasopressin and oxytocin. Moreover, this peptide increased additionally the release of both neurohypophysial hormones stimulated previously by peripheral osmotic challenge. On the other hand, the peptide increased the oxytocin but not vasopressin secretion brought about by an intracerebroventricular injection of hypertonic saline thus suggesting that the central osmotic stimulation decreases the sensitivity of vasopressin neurons to glucagon-like peptide-1 (7–36) amide. It is concluded that glucagon-like peptide-1 (7–36) amide may affect the secretory activity of the hypothalamo–neurohypophysial system under acute osmotic challenge.

HEPATOPROTECTOR PROPERTIES OF ERYXIN IN ANIMALS WITH ACUTE DRUG-INDUCED HEPATITIS MODEL

A considerable incidence of side effects caused by drug toxicity and the high lethal outcome of such cases make drug-induced hepatitis an important current problem [1]. Prophylaxis and elimination of liver disorders developed in the course of drug administration can be provided by widely used hepatoprotectors [2 – 4]. Unfortunately, the assortment of modern hepatoprotectors is very restricted [5]. In recent years, public health institutions of the Republic of Uzbekistan widely use the domestic drug eryxin (eriksin) developed at the Institute of the Chemistry of Plant Substances (Academy of Sciences of the Republic of Uzbekistan, Tashkent). This drug represents a highly purified, deprotonated 1% aqueous hydrolysate prepared from the biomass of snakes of the Eryx genus (sand boas). The parent substance contains a complex of biologically active low-molecular-weight compounds, peptides, free amino acids, trace elements (Ca, Mg, K, Fe, S), compounds containing SH groups, etc. [6]. Eryxin exhibits no antigen properties and is administered in the form of intravenous injections in a dose of 1–2m l (1 –2 ampules) per day or via dropper in a dose of 2 – 3 ampules dissolved in 250 ml of a 5% glucose solution or isotonic NaCl solution. Eryxin is administered in cases of immunodeficient states, chronic inflammatory disorders, rheumatological diseases, tuberculosis, brucellosis, acute obstructive or chronic bronchitis, dermatosclerosis, and for the treatment of neoplasms [6]. It should be noted that pathogenesis of acute drug-induced hepatitis always includes inflammatory-dystrophic and necrotic phenomena, with a monooxygenase system (representing predominantly the genetically determined isoforms of cytochrome P-450 [1, 7, 8]) involved in the biotransformation of xenobiotics [9, 10]. Taking this into account, investigation into the hepatoprotector properties of eryxin is a promising direction of research. In this context, we aimed at a comparative study of the effects of eryxin and legalon on the activity of liver monooxygenases and some biochemical characteristics of blood plasma in experimental animals with model acute drug-induced hepatitis. We selected legalon as the reference drug because this preparation possesses pronounced hepatoprotector activity and is widely used in hepatology for the treatment of both acute and chronic hepatitis of various genesis [2, 8, 10].

Characteristics of seed germination in five non-halophytic Chinese desert shrub species

The effects of temperature, NaCl, and polyethylene glycol (PEG)-6000 on the seed germination of five non-halophytic Chinese desert shrub species were investigated. The minimum temperature of germination was 10°C for all species, and the maximum temperature varied among species from 25 to 35°C. Isotonic solutions of NaCl and PEG caused different effects on seed germination in all five species. When seeds were moistened with −5·0 MPa NaCl solution for 5 days, the seeds of three species completely lost their germinability, while the seed germinability of the other two species was much less affected. These results are discussed with regard to the rehabilitation of desertified lands in China by seed dispersal.

Amiloride-sensitive signals and NaCl preference and appetite: a lick-rate analysis.

Rats prefer hypotonic and isotonic NaCl solutions to water in long-access drinking paradigms. To focus on the role of taste signals in NaCl preference, licking patterns of rats with 30-s exposure to NaCl solutions (0-0.5 M) were examined when they were either water deprived, sodium depleted, or not deprived (NaCl mixed in dilute sucrose). In all three conditions, rats displayed a preference for NaCl. The addition of 100 microM amiloride, a sodium channel blocker, to NaCl did not change rats' licking when they were sodium replete but dramatically reduced licking when they were deplete. Transection of the chorda tympani (CT) nerve, an afferent pathway for amiloride-sensitive Na(+) signals, had no effect on NaCl preference in nondeprived rats and only a modest effect on those that were Na(+) deplete. Amiloride was found to exert significant suppression of NaCl intake in Na(+)-depleted rats with transection of the CT, supporting the existence of other afferent pathways for transmission of amiloride-sensitive Na(+) signalling. Together, these studies argue for the involvement of different neural signalling mechanisms in NaCl preference in the presence and absence of explicit Na(+) need.

Water ingestion provides an early signal inhibiting osmotically stimulated vasopressin secretion in rats.

Dehydrated dogs are known to inhibit secretion of vasopressin (VP) within minutes after drinking water, before plasma osmolality (P(osmol)) diminishes. The present studies determined whether water ingestion causes a similar rapid inhibition of neurohypophyseal hormone secretion in rats. Adult rats were infused with 1 M NaCl (2 ml/h iv) for 240 min to stimulate VP and oxytocin (OT) secretion. After 220 min of infusion, rats were given water to drink for 5 min, and blood samples were taken 5 and 15 min later for RIA. Plasma VP (pVP) was much lower when rats ingested water than when they drank nothing even though P(osmol) was not significantly altered. Plasma OT (pOT) was affected similarly. In contrast, no effects on pVP or pOT occurred when rats drank isotonic NaCl solution for 5 min in amounts comparable to the water intakes (approximately 5.5 ml). These results suggest that neurohypophyseal secretion of VP and OT in rats is inhibited rapidly by water drinking, and that this inhibition is mediated by a visceral signal of osmotic dilution rather than by the act of drinking per se.

Mineralocorticoid receptor knockout mice: Lessons on Na+ metabolism

The mineralocorticoid receptor (MR) binds aldosterone and glucocorticoids with equal affinity. In aldosterone target tissues, like the epithelial cells of the distal colon and the principal cells of the collecting ducts in the kidney, the MR is protected from glucocorticoids by the action of the enzyme 11beta-hydroxysteroid-dehydrogenase type 2 (11betaOHSD2), allowing aldosterone to specifically activate the receptor. However, in MR-expressing cells, which lack 11betaOHSD2, like the neurons of the limbic system in the brain, MR is mainly activated by glucocorticoids. MR knockout mice die in the second week after birth, showing at day 8 symptoms of pseudohypoaldosteronism with hyponatremia, hyperkalemia, high renal salt wasting, and a strongly activated renin-angiotensin-aldosterone system (RAAS). The activity of the amiloride-sensitive epithelial Na+ channel (ENaC) is strongly reduced in colon and kidney, but there is no down-regulation of the mRNA abundance of the three ENaC subunits. Daily subcutaneous injections of isotonic NaCl solution until weaning and continued oral NaCl supply lead to survival of the MR knockout mice. The NaCl-rescued MR knockout mice display a strongly enhanced fractional renal excretion of Na+, hyperkalemia, and a persistently strongly activated RAAS. There is almost no renal ENaC activity. The renal mRNA abundance of alphaENaC is reduced by 30%, whereas betaENaC and gammaENaC are not altered.

Permeability Characteristics of Monovalent Cations in Atrial Myocytes of the Rat Heart

We investigated the permeability of Cs+ and Na+ through various ion channels in rat atrial myocytes using the whole-cell voltage-clamp technique. With isotonic CsCl (140 mM) on both sides of the membrane and nominally [Ca2+]o-free conditions, depolarising clamp pulses induced an increase of outward currents which showed a biphasic time course. Repolarisation to the holding potential induced inward tail currents. With isotonic NaCl, depolarisation also induced outward currents which showed a monotonic decay, but inward tail currents were not observed. Both in NaCl and CsCl, currents were hardly affected by TEA (10 mM), 4-AP (5 mM) and DIDS (100 microM). Nicardipine (1 M) almost completely blocked time-dependent outward currents in isotonic NaCl solution, leaving only time-independent currents which showed linear I-V relationship. In isotonic CsCl conditions, nicardipine blocked outward current considerably, but there still remained time-dependent outward currents and inward tail currents. Addition of E-4031 (2-20 M) which is known as a specific blocker of the rapidly activating delayed rectifier K+ current (IKr) completely blocked these time-dependent outward and inward currents, leaving only a time-independent current. Time-independent currents recorded in the presence of nicardipine and E-4031 were inhibited by GdCl3, which is known to block non-selective cation (NSC) currents. From these results, it was suggested that NSC current in atrial myocytes can be investigated in isotonic Cs+ or Na+ solution in the presence of Ca2+ channel and IKr blockers.

Nickel and chromium release from stents into isotonic NaCl solution investigated with the aid of neutron activation analysis

Stent placement has become an effective treatment for abrupt or threatened vessel closure in recent years. As a preliminary investigation on the allergy problem towards to Ni and Cr metals from implanted stent, the release of the two elements from stents into isotonic NaCl solution was investigated. Trace amounts of Ni and Cr metals released from stents are successfully determined by neutron activation analysis (NAA). An extreme experimental condition is applied in the NAA procedure to reach a low detection limit of 1 ng Ni. A conclusion is given about the influence on the Ni and Cr release from gold coating, from the expansion, and from the different incubation periods.

Adherent surface mucus gel restricts diffusion of macromolecules in rat duodenum in vivo.

The aim of this study was to investigate the permeability of the adherent mucus gel layer in rat duodenum in vivo to macromolecules applied in the lumen. Rats were anesthetized with thiobarbiturate, and the duodenum was perfused with isotonic NaCl solution containing large-molecular-size secretagogues. Effects on mucosal HCO(-)(3) secretion and blood-to-lumen (51)chromium-labeled EDTA clearance were used as indexes that compounds had migrated across the mucus layer. Exposure to a low concentration of papain (10 U/100 ml) for 30 min removed the mucus layer without damage to the epithelium and induced or markedly enhanced HCO(-)(3) secretory responses to cholera toxin (molecular mass of 85 kDa) or glucagon (3.5 kDa). Water extracts from a VacA cytotoxin (89 kDa) producing Helicobacter pylori strain, but not from a toxin-negative isogenic mutant, caused a small increase in HCO(-)(3) secretion but only after the mucus layer had been removed by papain. The duodenal surface mucus gel thus significantly restricts migration of macromolecules to the duodenal surface. Release of bacterial toxins at the cell-mucus interface may enhance or be a prerequisite for their effects on the gastrointestinal mucosa.

Thermal damage to the corneal endothelium in diode laser thermokeratoplasty

Laser thermokeratoplasty (LTK) can be applied for correction of hyperopia and astigmatism by means of concentrically placed coagulations. Because of the temperature rise during coagulation, damage of the endothelial cells directly below the irradiated areas is possible. In this study, we examined the extent of the thermally denatured zones for different laser parameters and the threshold of thermal endothelial damage as a function of of temperature and duration of elevated temperatures. The threshold for thermal damage of endothelial cells was determined in isotonic NaCl solution for temperature exposures of 10 s and 1 min in a water bath. To determine the damage zones, corneas were irradiated under standardized conditions with a continuously emitting infrared (cw-IR) laser diode at various wave-lengths and different power values and were stained after preparation with trypan blue and alizarine red. The extensions of the damage zones were compared with calculated isotherms. Fifty percent cell damage was found at temperatures of 65 degrees C for heating times of 10 s and at 59 degrees C for 1 min. With thicker corneas, less laser power and higher absorption coefficients, the damage zone was reduced. The damage range determined corresponded to the calculated isotherms of 60 degrees C and 70 degrees C. Regarding clinical LTK, a loss of endothelial cells can be predicted and minimized or totally avoided by choosing the appropriate irradiation parameters.

Cerium-doped diosmectite for topical application studies of the cerium–clay interaction

Cerium is used for its antiseptic and immunomodulatory properties in burn injury. We have developed a cerium-doped clay to replace existing ointments. Adsorption and release of cerium (Ce 3+) by diosmectite were studied at 22±2°C, in the presence of various other cationic species. Simple spectrofluorimetric determination of cerium was used ( λexc=240 nm/ λem=360 nm). Cerium binding reached a plateau within 2 min and was a function of the electrolyte content of the solution in contact with the clay. Langmuir isotherm treatment led to a maximal binding capacity of 66 mg of Ce 3+ per gram of clay. Partial release occurred within 2 min (19% in the presence of isotonic NaCl solution). The ionic strength of the solution, and the ionic radius and charge of the electrolytes present in the bathing solution significantly influenced cerium release, in contrast to pH and temperature changes. These results strongly point to a cationic exchange mechanism between diosmectite and cerium solution.

Pressor Responses to Intracerebroventricular Infusion of Hypertonic NaCl in Renal Hypertensive Rats

Background:The sodium concentration in the central nervous system may play an important role in cardio- vascular function and body fluid regulation. The purpose of this investigation was to examine the effects of intracerebroventricular (ICV infusion of hypertonic NaCl solutions on the cardiovascular responses in normotensive and 2-kidney, 1 clip (2K1C renal hypertensive rats. Methods:2K1C hypertension was made by clipping the left renal artery and were used 4 weeks later. Age-matched control rats received a sham treatment. Under thiopental (50 mg/kg, IP anesthesia, both isotonic and hypertonic NaCl solutions (0.15 M, 0.6 M and 1.2 M were ICV applied, while blood pressure and heart rate (HR responses were continuously monitored. Results:Central administration of hypertonic NaCl solution caused an elevation in mean arterial pressure (MAP and HR in both normotensive and 2K1C hypertensive rats. The response magnitude in the blood pressure was positively correlated to the NaCl concentration in normotensive rats, while the pressor responses to hypertonic NaCl were comparable regardless of the concentration of NaCl in hypertensive rats. Despite of the HR responses were similar in between two groups, the magnitude of the MAP increases were more elevated in hypertensive than in normotensive control rats. Isotonic NaCl solution, when centrally applied, caused an elevation in blood pressure only in hypertensive rats. Conclusion:These results indicate that the central sensitivity to sodium chloride is altered in 2K1C renal hypertensive rats. ( ( ( (Korean Circulation J 1999;29( ( ( (2 :216-221

The Influence of Cryopreservation on Murine Oocyte Water Permeability and Osmotically Inactive Volume

Osmotic experiments were performed on unfrozen (N= 18) and cryopreserved (N= 21) ICR murine oocytes in order to determine whether a standard cryopreservation process alters membrane water permeability (hydraulic conductivity,Lp) and/or osmotically inactive volume (Vb). Oocytes, initially in an isotonic (288 mOsm) NaCl solution, were exposed to 900 mOsm NaCl in a microdiffusion chamber. Cell size changes were videotaped and analyzed using a parameter estimation program. Best estimates for a two-parameter model (LpandVb) which includes the osmotically inactive volume as a fitting parameter are presented for the first time. The cryopreservation process produced no significant difference between the meanLpor the meanVbvalues for the unfrozen control population (Lp= 0.64 ± 0.15 μm/min/atm,Vb= 24.7 ± 2.9%) and the cryopreserved population (Lp= 0.63 ± 0.12 μm/min/atm,Vb= 28.0 ± 10.8%). While the cryopreservation process did not cause significant changes in the mean values ofLp,Vb, or the variability ofLp, it did produce more variability ofVb. The cause of the increased variability ofVbproduced by cryopreservation is unknown. These results suggest that the osmotic properties of unfrozen control oocytes can be used as a reasonable approximation for frozen–thawed oocytes. They also suggest that multiple parameter models and parameter estimation methods may be useful in developing a more comprehensive understanding of the more subtle alterations in osmotic properties that were detected here. Statistical tests were also used for the first time to confirm the assumption that all of the experimental populations were derived from normal distributions.

Amiloride inhibits taste nerve responses to NaCl and KCl in Sprague-Dawley and Fischer 344 rats

In a two-bottle test, Sprague-Dawley rats preferentially consume a greater amount of hypotonic and isotonic NaCl solutions relative to water, whereas inbred Fischer 344 (F344) rats fail to prefer NaCl solutions at any concentration relative to water. To determine whether taste contributes to this strain difference, we measured the integrated neural responses of the chorda tympani nerve to a concentration range of NaCl and KCl solutions. The amiloride-sensitive component of the taste nerve response was assessed by adding amiloride during salt stimulation in Experiment 1, and by pretreating the taste receptors with amiloride prior to salt stimulation in Experiment 2. Adding amiloride to NaCl during sustained neural activity suppressed chorda tympani nerve responses more than pretreating the tongue with amiloride. Adding amiloride during salt stimulation also partially suppressed chorda tympani neuron responses to KCl, a presumed control stimulus. The neural responses of the chorda tympani nerve to NaCl and KCl were similar for salt-avoiding F344 and salt-preferring Sprague-Dawley rats. However, amiloride pretreatment suppressed the taste nerve responses to NaCl significantly less in F344 rats than in Sprague-Dawley rats. The strain difference in the amiloride-sensitive component of the taste response may contribute to the difference in NaCl preference.