NaCl Preference Research Articles

Abstract P170: Beta-arrestin-2 Sustains Experience-dependent Plasticity Of Sodium Ingestive Behavior

Elevated sodium (Na) intake is a risk factor for hypertension. Na “appetite” is defined by the persistence of Na consumption despite adverse conditions and is commonly assessed through ingestion of aversive NaCl solutions. β-arrestins interact with many G Protein-Coupled Receptors (GPCR), including the Angiotensin II Type 1 Receptor (AT 1 R). β-arrestins play roles in many physiological processes including regulation of cardiovascular function, sensory processing, and energy metabolism. Recent work published by our group concluded that β-arrestin-2 deficient (Arrb2 KO ) mice exhibited increased ingestion of NaCl solutions even in the absence of physiological need. Thus, we hypothesize that β-arrestin-2 plays a role in experience-based Na seeking behavior and elevated NaCl consumption. To explore this question, mice were first presented with a 2-bottle choice of water and 0.15 NaCl.; then exposed to aversive 0.3 M NaCl exclusively for 24h; followed by the same 2-bottle choice as at baseline. Usually, this manipulation would yield a reduction in preference for previously preferred concentrations of NaCl, demonstrating experience-based plasticity of NaCl preference. At baseline, male Arrb2 KO mice exhibited the expected increase in preference for 0.15 M NaCl vs water (control n=14, 34±5%; Arrb2 KO n=10, 55±7%, p<0.01). Following 24h hypertonic (0.3 M) saline exposure, both groups displayed a decrease in 0.15 M NaCl solution intake and preference (days 1+2 after exposure: 10±2% vs 21±4%, p<0.01 vs baseline within each group, and p=0.15 between groups). While control mice maintained this significant suppression of preference for 0.15 M NaCl beyond one week after 24h 0.3 M NaCl exposure (e.g., days 7+8 after exposure: controls 22±3% p=0.01 vs baseline), preference of Arrb2 KO mice for 0.15 M NaCl returned to baseline levels within days (e.g., days 7+8 Arrb2 KO 45±9%, p=0.08 vs baseline, and p<0.01 vs control). This supports the idea that Arrb2 is implicated in Na ingestive behaviors, potentially influencing memory maintenance and retrieval. Future studies aim to characterize the role of Arrb2 downstream of the AT 1 R within specific brain regions in Na consummatory behaviors.

Abstract 104: DOCA Induced Natriorexigenic and Hypertensive Effects are Blunted by Central Infusion of AT1 Receptor Biased Agonist TRV120027

TRV120027 (TRV) is biased agonist at the angiotensin AT1 receptor that activates the β-arrestin pathway, causing receptor internalization. AT1 signaling within the hypothalamus is required for the blood pressure, polydipsia, and sodium appetite phenotypes in the deoxycorticosterone acetate (DOCA)-salt model of salt-sensitive hypertension (HTN). We hypothesized that TRV action within the hypothalamus would antagonize the effects of DOCA-salt. Using a novel luciferase-based HiBit tagging assay, we confirmed that TRV decreased AT1 surface density in immortalized mouse hypothalamic N43/5 cells by 35±4% compared to vehicle (n=4, p<0.05), and this was prevented by pretreatment with losartan. TRV (0.05 μg/h, icv n=5-10) or aCSF (n=4-8) were chronically infused into C57BL/6J mice implanted with DOCA (50 mg, sc) pellets, compared with mice undergoing sham surgeries (CTL, n=5-8), and animals were offered chow, water and NaCl ad libitum . DOCA increased fluid intake at baseline (CTL 4±0 vs 15±1 mL/day DOCA+aCSF, p<0.05), 0.15 mol/L NaCl (3.8±0 vs 20±2, p<0.05), 0.3 mol/L (4±0 vs 26±2, p<0.05), and 0.45 mol/L (4±0 vs 28±3, p<0.05). DOCA-induced polydipsia was not affected by TRV at baseline (DOCA+aCSF 19±1 vs 15±0 mL/day, p>0.05), 0.15 mol/L NaCl (27±4 vs 20±2 mL/day, p>0.05), 0.3 mol/L (25±4 vs 26±2 mL/day, p>0.05), or 0.45 mol/L (25±3 vs 28±3 mL/day, p>0.05). However, TRV prevented DOCA-induced increase of NaCl preference at NaCl 0.15 mol/L (DOCA+aCSF 29±1 vs 14±2% DOCA+TRV, p<0.05), 0.30 mol/L (29±3 vs 5±1%, p<0.05), and 0.45 mol/L (29±3 vs 5±1%, p<0.05). DOCA+aCSF exhibited increased heart weight / tibia length (CTL 8±0 vs 10±0 g/m DOCA+aCSF, p<0.05), which was prevented by TRV (8±0, p<0.05). In a separate cohort, DOCA-implanted mice were provided only 0.9% NaCl drink for one week, and mean arterial pressure (MAP) was recorded after acute injection of TRV (2 μg icv n=5) or aCSF (n=5). MAP was elevated with DOCA-salt (141±5 mmHg), and at 40 minutes post injection, TRV decreased MAP by 17±4 mmHg (p<0.05). These data underscore the potential for AT1 receptor-biased agonists as tools to treat subtypes of HTN characterized by salt sensitivity and low circulating renin activity.

Taste preference changes throughout different life stages in male rats.

Taste preference, a key component of food choice, changes with aging. However, it remains unclear how this occurs. To determine differences in taste preference between rats in different life stages, we examined the consumption of taste solutions and water using a two-bottle test. Male Sprague-Dawley rats of different ages were used: juvenile (3–6 weeks), young adult (8–11 weeks), adult (17–20 weeks), middle-aged (34–37 weeks), and old-aged (69–72 weeks). The intakes of the high and low concentration solutions presented simultaneously were measured. We observed that the old-aged group had lower preference ratios for 0.3 M sucrose and 0.1 M MSG in comparison with other groups. The preference ratio for 0.03 mM QHCl was higher in the middle-aged group than in the three younger groups and higher in the old-aged group than the juvenile group. The taste preferences for HCl and NaCl did not significantly differ among the age groups. The old-aged group tended to prefer high concentrations of sucrose, QHCl, NaCl, and MSG to low concentrations, indicating age-related decline in taste sensitivity. We also aimed to investigate differences between life stages in the electrophysiological responses of the chorda tympani nerve, one of the peripheral gustatory nerves, to taste stimuli. The electrophysiological recordings showed that aging did not alter the function of the chorda tympani nerve. This study showed that aging induced alterations in taste preference. It is likely that these alterations are a result of functional changes in other peripheral taste nerves, the gastrointestinal system, or the central nervous system.

Cyclic-AMP regulates postnatal development of neural and behavioral responses to NaCl in rats.

During postnatal development rats demonstrate an age-dependent increase in NaCl chorda tympani (CT) responses and the number of functional apical amiloride-sensitive epithelial Na+ channels (ENaCs) in salt sensing fungiform (FF) taste receptor cells (TRCs). Currently, the intracellular signals that regulate the postnatal development of salt taste have not been identified. We investigated the effect of cAMP, a downstream signal for arginine vasopressin (AVP) action, on the postnatal development of NaCl responses in 19–23 day old rats. ENaC-dependent NaCl CT responses were monitored after lingual application of 8-chlorophenylthio-cAMP (8-CPT-cAMP) under open-circuit conditions and under ±60 mV lingual voltage clamp. Behavioral responses were tested using 2 bottle/24h NaCl preference tests. The effect of [deamino-Cys1, D-Arg8]-vasopressin (dDAVP, a specific V2R agonist) was investigated on ENaC subunit trafficking in rat FF TRCs and on cAMP generation in cultured adult human FF taste cells (HBO cells). Our results show that in 19–23 day old rats, the ENaC-dependent maximum NaCl CT response was a saturating sigmoidal function of 8-CPT-cAMP concentration. 8-CPT-cAMP increased the voltage-sensitivity of the NaCl CT response and the apical Na+ response conductance. Intravenous injections of dDAVP increased ENaC expression and γ-ENaC trafficking from cytosolic compartment to the apical compartment in rat FF TRCs. In HBO cells dDAVP increased intracellular cAMP and cAMP increased trafficking of γ- and δ-ENaC from cytosolic compartment to the apical compartment 10 min post-cAMP treatment. Control 19–23 day old rats were indifferent to NaCl, but showed clear preference for appetitive NaCl concentrations after 8-CPT-cAMP treatment. Relative to adult rats, 14 day old rats demonstrated significantly less V2R antibody binding in circumvallate TRCs. We conclude that an age-dependent increase in V2R expression produces an AVP-induced incremental increase in cAMP that modulates the postnatal increase in TRC ENaC and the neural and behavioral responses to NaCl.

Ethanol Consumption in Mice Lacking CD14, TLR2, TLR4, or MyD88.

Molecular and behavioral studies support a role for innate immune proinflammatory pathways in mediating the effects of alcohol. Increased levels of Toll-like receptors (TLRs) have been observed in animal models of alcohol consumption and in human alcoholics, and many of these TLRs signal via the MyD88-dependent pathway. We hypothesized that this pathway is involved in alcohol drinking and examined some of its key signaling components. Different ethanol (EtOH)-drinking paradigms were studied in male and female control C57BL/6J mice versus mice lacking CD14, TLR2, TLR4 (C57BL/10ScN), or MyD88. We studied continuous and intermittent access 2-bottle choice (2BC) and 1-bottle and 2BC drinking-in-the-dark (DID) tests as well as preference for saccharin, quinine, and NaCl. In the 2BC continuous access test, EtOH intake decreased in male TLR2 knockout (KO) mice, and we previously reported reduced 2BC drinking in male and female CD14 KO mice. In the intermittent access 2BC test, EtOH intake decreased in CD14 KO male and female mice, whereas drinking increased in MyD88 KO male mice. In the 2BC-DID test, EtOH drinking decreased in male and female mice lacking TLR2, whereas drinking increased in MyD88 KO male mice. In the 1-bottle DID test, EtOH intake decreased in female TLR2 KO mice. TLR2 KO and CD14 KO mice did not differ in saccharin preference but showed reduced preference for NaCl. MyD88 KO mice showed a slight reduction in preference for saccharin. Deletion of key components of the MyD88-dependent pathway produced differential effects on EtOH intake by decreasing (TLR2 KO and CD14 KO) or increasing (MyD88 KO) drinking, while deletion of TLR4 had no effect. Some of the drinking effects depended on the sex of the mice and/or the EtOH-drinking model.

Fluid deprivation increases isotonic NaCl intake, but not hypertonic salt intake, under normal and heated conditions in obese Zucker rats

In the course of exposure to fluid deprivation and heated environment, mammals regulate their hydromineral balance and body temperature by a number of mechanisms including sweating, water and salt intakes. Here we challenged obese Zucker rats, known to have a predisposition to hypertension, with 0.9%NaCl alone or with 2%NaCl solution + water to drink under fluid deprivation and heated conditions. Food and fluid intakes, body weight, diuresis and natriuresis were measured daily throughout. Serum aldosterone levels and Na+ concentration were also analyzed. Data showed that obese and lean rats presented similar baseline measurements of food, 0.9%NaCl and fluid intakes, diuresis and fluid balance; whereas hypertonic 2%NaCl consumption was almost absent. Before and during fluid deprivation animals increased isotonic but not hypertonic NaCl intake; the obese showed significant increases in diuresis and Na+ excretion, whereas, total fluid intake was similar between groups. Heat increased isotonic NaCl intake and doubled natriuresis in obese which were wet on their fur and displayed a paradoxical increase of fluid gain. Fluid deprivation plus heat produced similar negative fluid balance in all groups. Body weight losses, food intake and diuresis reductions were amplified under the combined conditions. Animals exposed to 2%NaCl showed higher circulating levels of aldosterone and obese were lower than leans. In animals which drank 0.9%NaCl, obese showed higher serum levels of Na+ than leans. We conclude that in spite of their higher sensitivity to high salt and heat obese Zucker rats can control hydromineral balance in response to fluid deprivation and heat by adjusting isotonic NaCl preference with sodium balance and circulating levels of aldosterone. This suggests a key hormonal role in the mechanisms underlying thermoregulation, body fluid homeostasis and sodium intake.

Inter-generic and inter-habitat variation in the demand for sodium by Neotropical ants

Sodium is a potentially limiting element for ant populations. Here we examined the extent to which the demand for sodium varies among different ant genera and habitats. Sucrose and NaCl baits were placed on the ground and in trees located within three types of savanna and one type of forest. Tree-dwelling ants were found ca. two times more often on NaCl than on sugar baits, whereas on ground the frequency of ants on NaCl baits was about 20 % lower than on sugar baits. This difference seems to be explained by compositional differences between the tree- and ground-dwelling faunas. Camponotus and Cephalotes were much more frequent in trees than on ground, and both showed a preference for NaCl over sucrose baits. In contrast, the most frequent ants on the ground were Pheidole, and these were mainly attracted to sucrose baits. Genera-specific patterns of bait use did not differ significantly among habitats in spite of potential differences in the availability of sucrose (in honeydew and plant exudates) and sodium (in animal matter) for ants among these same habitats. In all habitats, genera that are predominantly carnivorous were comparatively less attracted to NaCl baits than those that rely more on plant matter. These findings lend additional support to the idea that sodium limitation is associated with the ant´s trophic position. At the community level, thus, the attractiveness of sodium sources to ants will depend both on rates of external sodium inputs as well on community composition.

Activation of the renin-angiotensin system, specifically in the subfornical organ is sufficient to induce fluid intake.

Increased activity of the renin-angiotensin system within the brain elevates fluid intake, blood pressure, and resting metabolic rate. Renin and angiotensinogen are coexpressed within the same cells of the subfornical organ, and the production and action of ANG II through the ANG II type 1 receptor in the subfornical organ (SFO) are necessary for fluid intake due to increased activity of the brain renin-angiotensin system. We generated an inducible model of ANG II production by breeding transgenic mice expressing human renin in neurons controlled by the synapsin promoter with transgenic mice containing a Cre-recombinase-inducible human angiotensinogen construct. Adenoviral delivery of Cre-recombinase causes SFO-selective induction of human angiotensinogen expression. Selective production of ANG II in the SFO results in increased water intake but did not change blood pressure or resting metabolic rate. The increase in water intake was ANG II type 1 receptor-dependent. When given a choice between water and 0.15 M NaCl, these mice increased total fluid and sodium, but not water, because of an increased preference for NaCl. When provided a choice between water and 0.3 M NaCl, the mice exhibited increased fluid, water, and sodium intake, but no change in preference for NaCl. The increase in fluid intake was blocked by an inhibitor of PKC, but not ERK, and was correlated with increased phosphorylated cyclic AMP response element binding protein in the subfornical organ. Thus, increased production and action of ANG II specifically in the subfornical organ are sufficient on their own to mediate an increase in drinking through PKC.

Dietary zinc status reversibly alters both the feeding behaviors of the rats and gene expression patterns in diencephalon

Nutritional status influences feeding behaviors, food preferences, and taste sensations. For example, zinc-deficient rats have been reported to show reduced and cyclic food intake patterns with increased preferences for NaCl. Although some impairments of the central nervous and endocrine systems have been speculated to be involved in these phenomena, the effects of short-term zinc deficiency on the brain have not been well examined to date. In this study, we performed a comprehensive analysis of the gene expression patterns in the rat diencephalon, which is a portion of the brain that includes the hypothalamus and thalamus, after short-term zinc deficiency and also during zinc recovery. The rats showed reduced and cyclic food intake patterns with increased salt preferences after a 10-day dietary zinc deficiency. A comparative analysis of their diencephalons using cDNA microarrays revealed that approximately 1% of the genes expressed in the diencephalons showed significantly altered expression levels. On the other hand, a 6-day zinc supplementation following the deprivation allowed for the recovery to initial food intake behaviors and salt preferences. The expression levels of most of the genes that had been altered by exposure to zinc deficient conditions were also recovered. These results show that feeding behaviors, taste preferences and gene expression patterns in the diencephalon respond quickly to changing zinc levels.

NaCl Taste Thresholds in 13 Inbred Mouse Strains

Molecular mechanisms of salty taste in mammals are not completely understood. We use genetic approaches to study these mechanisms. Previously, we developed a high-throughput procedure to measure NaCl taste thresholds, which involves conditioning mice to avoid LiCl and then examining avoidance of NaCl solutions presented in 48-h 2-bottle preference tests. Using this procedure, we measured NaCl taste thresholds of mice from 13 genealogically divergent inbred stains: 129P3/J, A/J, BALB/cByJ, C3H/HeJ, C57BL/6ByJ, C57BL/6J, CBA/J, CE/J, DBA/2J, FVB/NJ, NZB/BlNJ, PWK/PhJ, and SJL/J. We found substantial strain variation in NaCl taste thresholds: mice from the A/J and 129P3/J strains had high thresholds (were less sensitive), whereas mice from the BALB/cByJ, C57BL/6J, C57BL/6ByJ, CE/J, DBA/2J, NZB/BINJ, and SJL/J had low thresholds (were more sensitive). NaCl taste thresholds measured in this study did not significantly correlate with NaCl preferences or amiloride sensitivity of chorda tympani nerve responses to NaCl determined in the same strains in other studies. To examine whether strain differences in NaCl taste thresholds could have been affected by variation in learning ability or sensitivity to toxic effects of LiCl, we used the same method to measure citric acid taste thresholds in 4 inbred strains with large differences in NaCl taste thresholds but similar acid sensitivity in preference tests (129P3/J, A/J, C57BL/6J, and DBA/2J). Citric acid taste thresholds were similar in these 4 strains. This suggests that our technique measures taste quality-specific thresholds that are likely to represent differences in peripheral taste responsiveness. The strain differences in NaCl taste sensitivity found in this study provide a basis for genetic analysis of this phenotype.

Sweet and Salty Taste Responses in Alcohol‐preferring (P) and Alcohol‐Nonpreferring (NP) Rats

Naïve P rats showed increased mRNA and protein expression of T1R3 (a sweet taste receptor component) and TRPV1t (a putative benzamil (Bz)-insensitive salt taste receptor) in the anterior tongue and small intestine mucosal cells relative to NP rats. We investigated if up-regulation of receptors alter neural and behavior responses to sweet and salty stimuli in P rats. P rats showed a greater preference for 5% ethanol and 10% sucrose and greater chorda tympani (CT) taste nerve responses to sucrose, SC45647 and NaCl+Bz. Benzamil decreased NaCl preference in NP rats but not in P rats. A TRPV1t agonist, Maillard reacted peptides conjugated with galacturonic acid (GalA-MRPs), decreased preference for NaCl+Bz in P but not in NP rats. In P rats the concentration-CT response relation for ethanol and 0.5 M sucrose was higher and shifted to the left relative to NP rats. In naïve P rats and in NP rats chronically given 5% ethanol in a no choice paradigm, the concentration-CT response relation for TRPV1t agonists (resiniferatoxin and GalA-MRPs) and NaCl+Bz was enhanced and shifted to the left relative to naïve NP rats, but was unchanged in P rats. In NP rats, TRPV1t activity was increased by forced alcohol consumption to levels comparable to those of P rats. We conclude that chronic alcohol ingestion alters sweet and salt taste responses and may affect chemoreception in the gastrointestinal tract as well. Supported by DC-000122 and DC-005981.

The Brain Renin-Angiotensin System Controls Divergent Efferent Mechanisms to Regulate Fluid and Energy Balance

The renin-angiotensin system (RAS), in addition to its endocrine functions, plays a role within individual tissues such as the brain. The brain RAS is thought to control blood pressure through effects on fluid intake, vasopressin release, and sympathetic nerve activity (SNA), and may regulate metabolism through mechanisms which remain undefined. We used a double-transgenic mouse model that exhibits brain-specific RAS activity to examine mechanisms contributing to fluid and energy homeostasis. The mice exhibit high fluid turnover through increased adrenal steroids, which is corrected by adrenalectomy and attenuated by mineralocorticoid receptor blockade. They are also hyperphagic but lean because of a marked increase in body temperature and metabolic rate, mediated by increased SNA and suppression of the circulating RAS. β-adrenergic blockade or restoration of circulating angiotensin-II, but not adrenalectomy, normalized metabolic rate. Our data point to contrasting mechanisms by which the brain RAS regulates fluid intake and energy expenditure.

NaCl preference and water intake effects of food availability in median eminence polydipsia

Lesions in the median eminence (ME) produce a well-known neurological model of polydipsia and polyuria in rats. The effect of food availability ( ad lib/deprivation) on the polydipsic/polyuric behaviour of animals was tested. As expected, all lesioned rats developed strong polyuria and polydipsia during the first postoperative 24 h. This effect was maintained during day 2 when food was available ad lib (experiment A), but both polyuria and polydipsia were abolished when animals were deprived of food during the next 24 h (day 3). Animals deprived of food from the first post-operative day (experiment B) showed a significant reduction in the initial polyuria and polydipsia (day 1) on day 2, but these effects were again observed on day 3 when food was available ad lib. Finally, when food-deprived animals were able to choose between a 1.5% sodium chloride solution and water (experiment C), they preferentially chose (82% of total liquid consumed) the hypertonic saline solution (day 1); during the next 24 h (day 2), when only water was available, the polyuric/polydipsic effect was abolished but returned when food became available ad lib on day 3. Hence, the polyuria/polydipsia effect produced by ME lesions appears to be consistent during the first 24 h but might later be related to the availability of standard food and is completely abolished under food deprivation conditions. Preference for the hypertonic solution supports the volemic component of this syndrome and demonstrates the need for appropriate amounts of hypertonic nutrients to be consumed during the first 24 h.

Investigating the motivational mechanism of altered saline consumption following 5-HT1a manipulation.

The precise role played by serotonin (5-HT) in taste--an issue of great interest given the involvement of serotonin in human sensory and eating disorders--is a matter of considerable debate, perhaps because of the variety of methodologies that have been brought to bear by different researchers. Here, we use multiple methods to reveal the motivational mechanism whereby 5-HT(1A) receptor activation modulates drinking behavior. Subcutaneous injections of the selective 5-HT(1A) agonist 8-hydroxy-2-di-n-propylamino-tetralin (8-OH-DPAT), a drug that reduces 5-HT release by acting on presynaptic autoreceptors, dose-dependently increased consumption of 0.45 M NaCl in a one-bottle test. In a two-bottle test, however, 8-OH-DPAT-treated animals (30 microg/kg/ml) demonstrated decreased NaCl preference--although our detection of this effect was obscured by adaptation to the drug across days. Rats' performance in a brief access test confirmed that 8-OH-DPAT decreased preference for saline by both increasing water consumption and decreasing NaCl consumption. Finally, taste reactivity tests demonstrated that the latter result does not reflect decreased NaCl palatability. Overall, the results suggest that 8-OH-DPAT-induced 5-HT hypofunction increases thirst without substantially affecting the palatability of NaCl.

Food, water and NaCl consumption by 14 strains of rats.

We measured the voluntary intake of food, water, and NaCl by 8 male and 8 female rats from each of 3 outbred strains (Crl:CD(SD) IGS, LE, and Wistar) and 11 inbred strains (BN/SsNHsdMcwiCrl, BUF/CrCrl, COP/CrCrl, DA/OlaHsd, DahlSS/JrHsdMcwi, F344/NTac, FHH/EurMcwi, LEW/NHsd, Noble/CrCrl, PVG/OlaHsd, and SHR/NCrl). At 8-week old when tests began, there was a 182-g range in body weight (PVG-female=121±2 g, LE-male=303±4 g), a 17-g range in food intake (BN-female=11±1 g/d, SHR-male=28±2 g/d of AIN-76A diet), and a 34-mL range in water intake (BN-female=15±1 mL/d, FHH-male=49±4 mL/d of deionized water). In a series of 48-h two-bottle tests with a choice between water and progressively increasing concentrations of NaCl, all rat strains strongly preferred 3.16, 10, 31.6, and 100 mM NaCl relative to water. All except BUF, Noble and PVG preferred 178 mM NaCl relative to water. All except LEW, SHR, and DA avoided 316 mM NaCl, and all avoided 562 mM NaCl. There were few differences in NaCl preference between the sexes at any concentration. These results illustrate the substantial diversity of voluntary ingestive behavior in rats and set the stage for future genetic analyses.

Taste Solution Preferences of C57BL/6J and 129X1/SvJ Mice: Influence of Age, Sex, and Diet

To examine whether age influences taste solution preferences, we measured taste preferences of C57BL/6J and 129X1/SvJ mice given a series of 48-h 2-bottle tests with a choice between water and one of the following taste solutions: 2 mM saccharin, 5 mM citric acid, 30 microM quinine hydrochloride, 75 mM sodium chloride (NaCl), 10 mM inosine monophosphate (IMP), 50 mM calcium chloride (CaCl(2)), and 10% ethanol. We tested separate groups of male mice fed Teklad 8604 chow at ages 4, 6, 9, 12, 15, 20, 25, 30, 40, and 50 weeks and retested some of these mice at 54, 75, and 100 weeks and again at 125 weeks. Female mice fed chow were tested at ages 4, 12, 25, and 50 weeks and retested at 54, 75, 100, and 125 weeks. Male mice fed AIN-93G semisynthetic diet were tested at ages 4, 12, 25, and 50 weeks and retested at 54, 75, and 100 weeks. Concentration-response functions for each taste solution were collected from male and female mice fed chow aged 8 or 125 weeks. In general, the results showed that age had little effect on taste preferences. Exceptions included 1) a small increase in quinine hydrochloride preference between 54 and 125 weeks in mice of both strains and sexes, 2) a marked increase in NaCl preference between 4 and 12 weeks in female B6 mice, 3) a gradual decrease in IMP preference between 4 and 125 weeks in male and female 129 mice, 4) a marked decrease in CaCl(2) preference between 54 and 125 weeks in male and female 129 mice, and 5) a marked reduction in ethanol preference between 4 and 12 weeks in male B6 mice fed AIN-93G diet but not chow. These results show that over a wide range and with the exceptions noted, age contributes little to the variation in taste preferences observed in C57BL/6J and 129X1/SvJ mice.

Taste and acceptance of pyrophosphates by rats and mice

The palatability and taste quality of pyrophosphates were evaluated in a series of behavioral and electrophysiological experiments. In two-bottle choice tests with water, rats strongly preferred some concentrations of Na3HP2O7 and Na4P2O7, moderately preferred some concentrations of K4P2O7 and Fe4(P2O7)3, and were indifferent to or avoided all concentrations of Ca2P2O7 and Na2H2P2O7. The contribution of sodium to the preference for sodium pyrophosphates was ascertained: 1) Rats with a choice between Na4P2O7 and NaCl preferred 1 mM Na4P2O7 to 4 mM NaCl but preferred 40 or 150 mM NaCl to 10 mM Na4P2O7, 2) blocking salt taste transduction by mixing Na4P2O7 with amiloride reduced preferences but did not eliminate them, and 3) three mouse strains (FVB/J, C57BL/6J, and CBA/J) known to differ in sodium preference had the same rank order of preferences for Na3HP2O7 and NaCl, but peak preferences were higher for Na3HP2O7 than for NaCl. The taste qualities of pyrophosphates were determined by measuring taste-evoked responses of neurons in the nucleus of the solitary tract of rats. Across-neuron patterns of activity for sodium pyrophosphates were similar to that of NaCl but the pattern of Na3HP2O7 plus amiloride was unique from those of sweet, salty, sour, bitter, and umami stimuli. Taken together, the results indicate that the high palatability of some concentrations of Na3HP2O7 and Na4P2O7 is due partially to their salty taste, but there must also be another cause, which may include a novel orosensory component distinct from the five major taste qualities.

Detection of NaCl and KCl in TRPV1 Knockout Mice

Both amiloride-sensitive and -insensitive mechanisms contribute to NaCl taste transduction. The amiloride-sensitive mechanism relies on the epithelial Na(+) channel ENaC, which is widely expressed on the apical membrane of fungiform taste cells. The amiloride-insensitive mechanism, which predominates in circumvallate and foliate taste buds, was recently reported to involve a variant of the nonselective cation channel TRPV1. We performed 2-bottle preference and threshold experiments with TRPV1 knockout mice and wild-type (C57BL/6J) controls to test for NaCl preference and detection thresholds in the presence and absence of amiloride. Surprisingly, TRPV1 knockout mice not only detected NaCl in the presence of amiloride but they preferred NaCl over water at concentrations avoided by the wild-type mice. NaCl detection thresholds were between 2 and 3 mM for both genotypes. Amiloride increased the detection thresholds of wild-type mice but not knockout mice. The knockout mice also preferred 100 mM KCl compared with wild-type controls, suggesting that TRPV1 receptors may mediate a general aversive response to salts. Analyses of consumption data also revealed that TRPV1 knockout mice ingested more of the NaCl, with and without amiloride, and KCl solutions than the wild-type mice. However, comparisons of preference ratios and consumption volumes indicated that both wild-type and TRPV1 knockout mice avoided citric acid in quite a similar manner, suggesting that TRPV1 receptors do not mediate the detection of citric acid. These data, taken together, suggest that additional mechanisms must contribute to the amiloride-insensitive NaCl response.

Sex differences in electrophysiological and behavioral responses to NaCl taste.

We tested the hypothesis that sex differences in preference for NaCl are attributable to estrogen-mediated alterations in gustatory processing. Electrophysiological responses of the chorda tympani nerve to NaCl were blunted by estrogen treatment in ovariectomized female rats, suggesting that females are less sensitive to concentrated NaCl solutions during high estrogen conditions. In contrast, after a taste aversion was conditioned to 150-mM NaCl, estrogen- and oil-treated ovariectomized rats generalized the aversion to a lower concentration of NaCl than did males, suggesting that females are more sensitive to the taste of dilute NaCl solutions regardless of estrogen. Thus, sex differences in NaCl preferences may be attributable to differences in NaCl taste processing that involve both acute and developmental effects of estrogen.

Sodium preference and appetite in rats in an operant protocol

An operant task that may simulate natural foraging was used to study in Sprague–Dawley rats the need-free choice between water and various concentrations of NaCl solution, and the induction of sodium appetite during chronic deoxycorticosterone acetate (DOCA) treatment. The task imposed a fixed ratio foraging or procurement cost for access to each fluid. The relative preference for NaCl over water when both had the same procurement cost was similar to the preference/aversion function that has been reported for this strain of rat under free drinking conditions. As the procurement cost increased, the number of bouts taken per day of both water and NaCl decreased and the mean bout size increased. Administration of DOCA produced an increase in NaCl intake under all conditions, and this appetite was manifest as discrete episodes whose frequency and size were inversely related. Natural observations of sodium appetite in mammals, such as excursions to, often remote, mineral deposits, imply an episodic organization of this behavior. This report is the first laboratory simulation of environmental factors that affect the episodic or temporal organization of sodium appetite.