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Am J Physiol Regul Integr Comp Physiol 290: R1736-R1741, 2006. First published January 26, 2006; doi:10.1152/ajpregu.00043.2005
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WATER AND ELECTROLYTE HOMEOSTASIS

Antidipsogenic effects of a TRPV4 agonist, 4{alpha}-phorbol 12,13-didecanoate, injected into the cerebroventricle

Hiromi Tsushima and Mayumi Mori

Department of Cellular and Molecular Pharmacology, Nagoya City University Graduate School of Medical Sciences, Mizuho-ku, Nagoya, Japan

Submitted 21 January 2005 ; accepted in final form 23 January 2006


   ABSTRACT
TOP
 ABSTRACT
MATERIALS AND METHODS
 RESULTS
 DISCUSSION
 GRANTS
 REFERENCES
 
Transient receptor potential vanilloid 4 (TRPV4) is one member of the TRP superfamily of nonselective cation channels. Recently, the possibility has been raised that TRPV4 is an osmoreceptor, because it is found in the circumventricular organs where osmoreceptors are supposed to be distributed and because it is sensitive to osmotic pressure in in vitro experiments. In addition, TRPV4 knockout mice have abnormal osmosensitivity. In this study, effects of 4{alpha}-phorbol 12,13-didecanoate (4{alpha}-PDD), a TRPV4 agonist, on drinking behavior were examined to investigate roles for TRPV4 as an osmoreceptor in vivo in wild-type animals. Intracerebroventricular injections of 4{alpha}-PDD inhibited water intake under normal conditions in both light and dark periods of the day, after food deprivation, and after administration of angiotensin II. However, this drug did not influence increased water intake after administration of a hypertonic solution or after water deprivation that significantly increased plasma osmolality. Locomotor activity of the 4{alpha}-PDD-injected group decreased slightly compared with that of the vehicle-injected group; however, sweet taste, food intake, and body temperature were not different between the two groups. The antidipsogenic effects of 4{alpha}-PDD were blocked by preinjection into the ventricle of TRPV4 antagonists such as ruthenium red or gadolinium. These findings suggest that TRPV4 regulates drinking behavior under certain conditions, and the regulation interacts with the angiotensin II pathway.

water intake; hypothalamus; osmoreceptor; transient receptor potential vanilloid 4

HOMEOSTASIS OF OSMOTIC PRESSURES of body fluids is maintained on the basis of a balance between water intake and excretion. Osmoreceptors in the central nervous system (CNS) perform a key step in osmoregulation, a step that detects changes in osmotic pressures of body fluids. Osmoreceptors are suggested to be distributed in the circumventricular organs, especially the anterior wall adjacent to the third ventricle, the hypothalamic paraventricular and supraoptic nuclei where the cell bodies of vasopressin-containing neurons localize, and the subfornical organ (3). However, molecular identification of the osmoreceptor is not clarified.

The transient receptor potential (TRP) superfamily of nonselective cation channels are known to be present in many species from worms to man (5). Of six subfamilies, some members of the TRPV and TRPM subfamilies are temperature sensitive, although each has a different range of temperature sensitivity. Therefore, it is suggested that TRP channels of the CNS are involved in the regulation of body temperature, whereas those of the skin are involved in the regulation of thermosensation (17). Recently, Liedtke et al. (12) found that TRPV4, one member of the TRPV subfamily, is sensitive to osmotic pressure, as well as to temperature. TRPV4 is activated by hypotonic stimuli, resulting in an increase in intracellular Ca2+ concentration ([Ca2+]i). In addition, this channel is found in the circumventricular organ of the CNS (7, 12). Therefore, it has been suggested that TRPV4 functions as an osmoreceptor in the CNS. Since then, many studies have been published concerning TRPV4, and most of them were carried out using an electrophysiological technique or [Ca2+]i measurement method in TRPV4-transfected cells.

In 2003, two laboratories succeeded in generating TRPV4 knockout (KO) mice and showed that they have abnormal osmosensitivity (13, 14). Mizuno et al. (14) demonstrated that the urinary volume and ion excretion, water intake, and vasopressin release of the KO mice are normal under basal conditions, although hypertonic stimuli cause an increased release of vasopressin in the KO mice compared with wild-type (WT) mice. However, the KO mice are supposed to show some abnormality even under basal conditions if, as they point out, TRPV4 is constitutively activated by body temperature. In addition, it remains unexplained why KO and WT mice show similar responses to hypotonic stimuli. On the other hand, KO mice from the laboratory of Liedtke and Friedman (13) have shown abnormalities in plasma osmolality, water intake, and/or vasopressin release compared with WT mice under basal conditions and after both hyper- and hyposmotic stimuli. Concerning vasopressin release, the two kinds of KO mice show opposite responses to hypertonic stimuli. These discrepancies make it difficult to understand a role of TRPV4 as an osmoreceptor in the KO mouse phenotype. Therefore, it is necessary to investigate the role of TRPV4 as an osmoreceptor in WT animals. In this study, we examined the effect of the TRPV4 agonist 4{alpha}-phorbol 12,13-didecanoate (4{alpha}-PDD) (25), microinjected into the lateral ventricle, on the water intake of WT animals.


   MATERIALS AND METHODS
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 ABSTRACT
 MATERIALS AND METHODS
RESULTS
 DISCUSSION
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Animals and surgical procedure. Experiments were carried out according to the Guidelines for Animal Care and Use of Nagoya City University. The experimental protocol was approved by the Institutional Animal Care and Use Committee of Nagoya City University Graduate School of Medical Sciences. The detailed procedure is described elsewhere (21–23). We used male Wistar rats (9 wk old; SLC, Hamamatsu, Japan), which were housed in the animal center of Nagoya City University under controlled conditions (temperature, 23 ± 1°C; humidity, 65%; and a 12:12-h light-dark cycle with lights on at 0800 and off at 2000). After the rats were anesthetized with pentobarbiturate (40 mg/kg body wt ip), a guide cannula (AG-8; Eicom, Tokyo, Japan) was implanted into the right lateral ventricle and fixed to the skull with dental cement and small screws according to coordinates provided by Konig and Klippel's atlas (11). During the 1-wk postoperative recovery period, rats were acclimated to handling and to the experimental cage used for drug administration. We cannulated a total of 140 rats, 18 of which were removed from the study because of weight loss in 6 rats after the operation and because of cannula misplacement in 12 rats.

Measurements of water and food intakes. Measurements were carried out under one of the experimental conditions shown in Table 1: 1) food deprivation (food removed on the previous day; no access for 16 h); 2) normal conditions during light period without treatment; 3) normal conditions during dark period without treatment; 4) water deprivation (drinking water removed on the previous day; no access for 16 h); 5) hypertonic NaCl administration (intragastric administration of 4 ml of 0.5 M NaCl carried out just before the rat was returned to its home cage; free access to food and drinking water); and 6) sweet drinking (8% sucrose given to rats for 3 h from 1200 to 1500 for 4 days, and intakes of sucrose solution were measured at 1500). Drug was administered at 1150 on the fourth day. On the day of the experiment, drugs were microinjected into the lateral ventricle by using a microinjection cannula inserted into the guide cannula. The microinjection cannula was connected via a polyethylene tube to a microsyringe containing two drug solutions. These solutions were separated by a small air bubble. In addition, 1 µl of sterilized saline was loaded between the solutions with air bubbles. The total volume of the drug solutions was 10–13 µl. The interval between the first and second drug microinjections was 30 min. Five minutes after the second injection, the microinjection cannula was switched to a dummy cannula, and the rats were returned to their home cages. Water and food intakes were measured 0.5, 1.0, 1.5, 2.0, 3.0, and 4.0 h after administration of the second drug. In one animal, two measurements were carried out with an interval of at least 1 wk in one experimental series. The drug combinations were randomly selected. However, when effects of 4{alpha}-PDD on ANG II-induced drinking behavior was examined under normal conditions (Table 1, group 2b), three drug combinations were administered in one animal with an interval of 1 wk between them and in the following order: ANG II + vehicle, ANG II + 4{alpha}-PDD, and ANG II + vehicle.


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  		Table 1. Summary of experimental conditions and protocols for water intake measurement

 
Measurement of body temperature. When the guide cannula was implanted into the ventricle, a telemetry transmitter (Minimitter; Eicom) was embedded in the abdomen. Body temperature was measured 3 h after drug administration under the condition of group 2a in Table 1. In one animal, vehicle or 4{alpha}-PDD was randomly microinjected into the ventricle with an interval of at least 1 wk between injections.

Measurement of locomotor activity. Locomotor activity was measured as the number of sensor beam interruptions using an activity monitor (Automex II; Columbus Instruments, Columbus, OH) under the condition of group 2a in Table 1. In one animal, vehicle and 4{alpha}-PDD were randomly microinjected into the ventricle with an interval of at least 1 wk.

Measurement of plasma osmolality. Plasma osmolality was measured just after vehicle injection under the conditions of groups 1 and 4 in Table 1, 5–10 min after vehicle injection and free access to food under the conditions of group 1 (after food intake without water intake), 0.5 h after vehicle injection under the conditions of groups 2 and 5, and 2 h after vehicle injection under the condition of group 3 in Table 1. After decapitation of the rat, blood was collected into an ice-cold tube containing heparin and centrifuged at 3,000 rpm and 4°C. Plasma osmolality was measured using the freezing-point depression method (Osmostat OM-6020; Kyoto Daiichi Kagaku, Kyoto, Japan).

Statistics. All of the data are expressed as means ± SE. Statistical analyses were carried out using one-way or two-way ANOVA followed by Fisher's protected least significant difference (PLSD) test (post hoc test) or Student's t-test when appropriate (StatView 4.0; Macintosh). Differences were considered significant if the P value was <0.05.

Drugs. 4{alpha}-PDD was obtained from Alexis Biochemicals (San Diego, CA); ruthenium red (RR) was obtained from Latoxan (Valence, France); and gadolinium (Gad) and ANG II were obtained from Sigma (St. Louis, MO). The chemicals used were the highest grade. 4{alpha}-PDD was dissolved in DMSO and diluted with sterile physiological saline (Otsuka Chemicals, Tokyo, Japan) as used. The final concentration of DMSO was 8%. The other drugs were dissolved in sterile physiological saline.


   RESULTS
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 ABSTRACT
 MATERIALS AND METHODS
 RESULTS
DISCUSSION
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Effects of 4{alpha}-PDD on water intake. Food-deprived rats drink more water than normally fed rats once the food supply is resumed after food deprivation. Intracerebroventricular (ICV) injections of 4{alpha}-PDD (20 µg) significantly inhibited this increased water intake, although food intake was not influenced (Fig. 1, A–C). Water and food intakes under food deprivation conditions in Fig. 1 were simultaneously measured in the same animals. The 4{alpha}-PDD-induced antidipsogenic effect was dose dependent [water intakes for 0–0.5 h after injection: vehicle, 4.0 ± 0.4 g, n = 9; 20 µg 4{alpha}-PDD, 1.1 ± 0.4 g, n = 8; 10 µg 4{alpha}-PDD, 2.6 ± 0.3 g, n = 4; 2 µg 4{alpha}-PDD, 4.4 ± 1.4 g, n = 5; P < 0.05, 20 µg 4{alpha}-PDD vs. vehicle or 2 µg 4{alpha}-PDD groups by Fisher's PLSD test after one-way ANOVA: F(3, 22) = 4.12, P < 0.05]. Pretreatments with RR (1 µg) and Gad (1 µg), two TRPV4 antagonists (8), injected into the ventricle completely blocked 4{alpha}-PDD-induced effects (Fig. 1, A and B). ICV injection of 1 µg of either RR or Gad into the ventricle did not influence food and water intakes (data not shown). However, at ~5 min after administration of RR, the bodies of 2 of 20 rats shook a few times, and locomotor activity seemed to decrease. These rats recovered after 30 min and took in the same volume of food and water as the vehicle-injected rats.


Figure 1
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  		Fig. 1. Effects of 4{alpha}-phorbol 12,13-didecanoate (4{alpha}-PDD) on increased water (cumulative data, A; noncumulative data, B) and food intakes (C) in the same animals after a 16-h food deprivation [vehicle + vehicle, n = 9; vehicle + 4{alpha}-PDD, n = 8; ruthenium red (RR) + 4{alpha}-PDD, n = 6; gadolinium (Gad) + 4{alpha}-PDD, n = 6] and effects of 4{alpha}-PDD on water intake during the light (cumulative data, D; noncumulative data, E; vehicle, n = 9; 4{alpha}-PDD, n = 6) and dark periods of the cycle (F, n = 6). Results are expressed as means ± SE. There was a significant effect of treatment [F(3, 50) = 3.46, P < 0.05] and a significant interaction of treatment x time [F(3, 50) = 3.88, P < 0.05] in B; a significant effect of treatment [F(1, 52) = 9.86, P < 0.05] in E by two-way ANOVA; and a significant effect of treatment [F(2,15) = 5.00, P < 0.05] in F by one-way ANOVA. Post hoc tests (Fisher's protected least significant difference test) showed significant decreases in water intake after administration of 4{alpha}-PDD and significant inhibitions on the 4{alpha}-PDD-induced effects by RR and Gad in B, E, and F. Noncumulative values were used for the analysis. Measurements in A–C, in D and E, and in F were made under the conditions of groups 1, 2a, and 3, as shown in Table 1, respectively. In 1 animal, 2 measurements were carried out with an interval of at least 1 wk in 1 experimental series. The drug combinations were randomly selected. 4{alpha}-PDD was microinjected into the lateral ventricle at time 0. Pretreatment with vehicle/antagonist into the ventricle was carried out 0.5 h before 4{alpha}-PDD administration. 4{alpha}-PDD (20 µg) only inhibited increased water intake, not food intake after food deprivation. Also, 4{alpha}-PDD reduced water intake during the light and dark periods under the normal conditions without any treatment. These effects were blocked by pretreatment with 1 µg of RR or Gad, TRPV4 antagonists. RR or Gad alone did not influence food and water intakes (data not shown). We used 20 and 18 rats for experiments under food deprivation and normal conditions, respectively (including experiments for administration of RR or Gad alone, which are not shown in Fig. 1).

 
The antidipsogenic effects induced by 4{alpha}-PDD also were observed under the normal light-dark condition without any treatment (Fig. 1, D–F). The 4{alpha}-PDD-induced effect during the dark period was blocked by RR (1 µg) (Fig. 1F). Food intake during the dark period after 4{alpha}-PDD administration did not change compared with that after vehicle administration (4{alpha}-PDD: 14.8 ± 0.9 g, n = 6; vehicle: 16.0 ± 0.8 g, n = 6). RR itself did not influence water or food intakes during the dark period.

ANG II is well known to be one of the most powerful dipsogenic agents. Increased water intake after ICV injection of ANG II (100 ng) was completely inhibited by 4{alpha}-PDD in five of nine rats (Fig. 2). After 1 wk, when ANG II alone was injected into the same rats once more, this induced the dipsogenic effects to the degree similar to the first effects.


Figure 2
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  		Fig. 2. Effects of 4{alpha}-PDD on increased water intake after ANG II administration. Results are expressed as means ± SE (open bar, vehicle + vehicle, n = 9; hatched bar, vehicle + 4{alpha}-PDD, n = 8; shaded bar, ANG II + vehicle, n = 5; solid bar, ANG II + 4{alpha}-PDD, n = 5). *P < 0.05 vs. ANG II + vehicle (Student's t-test). Drugs were microinjected into the ventricle of the rat in the order of ANG II + vehicle, ANG II + 4{alpha}-PDD, and then ANG II + vehicle during the light period under the normal condition (group 2b in Table 1). The data for vehicle + vehicle and vehicle + 4{alpha}-PDD are from Fig. 1D. The inhibition by 4{alpha}-PDD of the ANG II-induced dipsogenic effects were observed in 5 of 9 rats. The effects after the first and third injection of ANG II in the same animal were not different statistically (data not shown).

 
After a 16-h deprivation of water or gastric administration of hypertonic NaCl solution, animals increased their ingestion of water. However, water intakes for 4 h after injection were not different between the 4{alpha}-PDD- and the vehicle-injected groups (Fig. 3).


Figure 3
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  		Fig. 3. Effects of 4{alpha}-PDD on increased water intake after a 16-h water deprivation (A) and intragastric administration of hypertonic NaCl (B). Results are expressed as means ± SE (water deprivation, n = 4; hypertonic NaCl, n = 5). The drug injected (vehicle or 4{alpha}-PDD) was randomly selected. In A, drinking water was removed for 16 h from the previous day (group 4 in Table 1). In B, the rats were orally administered 4 ml of 0.5 M NaCl via a rubber tube (group 5 in Table 1). The increased water intakes after hypertonic stimuli gradually reverted to normal values 1.5–2.0 h after administration. Under these conditions, 4{alpha}-PDD did not change water intake increased by hypertonic stimuli. Noncumulative values were used for the analyses.

 
Effects of 4{alpha}-PDD on other behavior. Because drinking behavior also is influenced by various factors such as locomotor activity, taste aversion, and body temperature, these factors were examined after 4{alpha}-PDD administration (20 µg). Locomotor activity was slightly, but significantly, decreased after administration of 4{alpha}-PDD compared with after administration of vehicle (Table 2). To examine changes in taste aversion or illness/malaise after 4{alpha}-PDD administration, we measured how much sucrose solution rats drank. As shown in Fig. 4A, ICV injections of 4{alpha}-PDD did not influence the intake of sucrose solution. Neither body temperature after 4{alpha}-PDD administration was different from those after vehicle administration (Fig. 4B).


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  		Table 2. Locomotor activity after ICV injection of 4{alpha}-PDD

 

Figure 4
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  		Fig. 4. Effects of 4{alpha}-PDD on sucrose intake (A) and body temperature (B). A: 8% sucrose solution was given from 1200 to 1500 for 4 days. Vehicle or 4{alpha}-PDD was administered at 1150 on the 4th day (n = 4, group 6 in Table 1). B: basal body temperature was 37.5 ± 0.3°C for the 4{alpha}-PDD-injected group and 37.5 ± 0.4°C for the vehicle-injected group (n = 5). The experimental conditions were the same as those for group 2a in Table 1. All results are expressed as means ± SE (sweet taste, n = 8; body temperature, n = 5). The drug injected (vehicle or 4{alpha}-PDD) was randomly selected. 4{alpha}-PDD did not change sucrose intake or body temperature, which influence water intake.

 
Plasma osmolality. Table 3 shows plasma osmolality under experimental conditions for water intake measurement. Osmolality increased after a 16-h water deprivation period and after gastric administration of hypertonic solution, when water intake was not changed by 4{alpha}-PDD (Fig. 3). On the other hand, under a 16-h food deprivation, the normal conditions, and an ANG II-pretreated condition, the antidipsogenic effect of 4{alpha}-PDD was shown and osmotic pressure did not change. Plasma osmolality under 16-h food deprivation condition tended to increase after some food intake (1.4 ± 0.3 g, n = 8) without water intake (5–10 min after the rat was given free access to food); however, this was not statistically significant compared with osmolality during light and dark periods.


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  		Table 3. Plasma osmolality under various conditions

 

   DISCUSSION
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 ABSTRACT
 MATERIALS AND METHODS
 RESULTS
 DISCUSSION
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We found that 4{alpha}-PDD microinjected into the lateral ventricle inhibits water intake under certain conditions such as normal conditions during light and dark periods, after food deprivation, and during an ANG II-induced thirst condition. Sweet taste, food intake, and body temperature did not differ between the 4{alpha}-PDD- and vehicle-injected groups. Locomotor activity slightly decreased after 4{alpha}-PDD administration; however, this decrease is probably not the cause of the antidipsogenic effect. Because food and sucrose intakes were not influenced by 4{alpha}-PDD administration despite a decrease in locomotor activity, the 4{alpha}-PDD-administered animals did not show taste aversion or illness/malaise. Therefore, it is presumed that the 4{alpha}-PDD-induced antidipsogenic effect is not a secondary effect. In addition, the effects were abolished by pretreatment with RR and Gad. Because 4{alpha}-PDD is an agonist of TRPV4 and RR and Gad are antagonists of TRPV4, these findings show that the antidipsogenic effect involves TRPV4.

TRPV4 was shown to be sensitive to hyposmotic stimuli in an in vitro experimental system and to be present in the osmoreceptor-containing circumventricular region by immunohistochemical studies (12). Antidipsogenic effects of 4{alpha}-PDD were not observed under any in vivo condition. Indeed, 4{alpha}-PDD did not inhibit increased water intake following water deprivation or intragastric administration of hypertonic solution. These conditions significantly increased plasma osmotic pressure. We hypothesized that neurons expressing TRPV4 detect decreased osmotic pressure and regulate water balance under physiological conditions in which animals detect as little as a 1–2% change from normal osmolality; this results in thirst and water intake (1). TRPV4 in the CNS may play an important role as an osmoreceptor to detect hyposmolality and to stop water intake under physiological conditions.

Large changes in plasma osmolality influence a variety of factors, including 1) ion/substrate concentration, 2) cell volume, 3) membrane stretch strength, and 4) membrane structure/composition (10, 24, 27), which are followed by activation of intracellular signaling, and ion channels, among others. The Nax channel was found to be a Na+ sensor in the CNS and influences drinking behavior (9). Bourque and colleagues (24, 27) demonstrated that osmotic changes affect the length of the cell membranes, cell volume, and activity of ion channels and then promote vasopressin release, resulting in the regulation of osmolality of body fluids. Activation of mitogen-activated protein kinases is well known under hypertonic conditions (2, 28). Therefore, multiple mechanisms may operate powerfully during large osmotic pressure changes. We suppose that the effect of TRPV4 is masked or that TRPV4 is not involved in the regulation under these conditions.

Liedtke et al. (13) demonstrated that KO mice decrease drinking water volume after hypertonic NaCl stimuli, compared with WT mice. This finding suggests that WT mice drink water as mediated through activation of TRPV4 under hypertonic conditions, although under hypotonic conditions, TRPV4 is activated and WT mice do not drink water. They proposed a hypothesis that there is another molecule presented in vivo that responds to hypertonic stimuli, as well as to hypotonic stimuli. This phenomenon is inconsistent with our result that the TRPV4 agonist did not affect water intake after hypertonic NaCl stimuli. The presumed reasons are as follows. 1) Because TRPV4 in WT animals is fully activated by hypertonic stimuli, agonist administration will not be able to produce an additional effect any longer. 2) KO mice are not only WT mice without TRPV4; the regulatory mechanisms in KO mice are considerably disturbed directly and indirectly by the absence of TRPV4 because they become adapted to this condition since embryos. Two kinds of KO mouse phenotypes reported from the laboratories of Lietdke et al. (13) and Mizuno et al. (14) are different from each other. 3) From the whole body of KO mice, all of the target molecules are deficient; therefore, there may be a discrepancy between the KO mouse phenotype and an agonist/antagonist-induced effect in WT mice after the drug affects limited sites.

Vasopressin release is also regulated by plasma osmolality. Vasopressin controls urine volume, which is the largest component of water excretion from the body, and plays an important role in the regulation of body fluid osmolality. The presence of osmoreceptors on the cell bodies of vasopressin-containing neurons is well known (3, 24). Electrophysiological studies demonstrated that they are nonselective cation channels and are activated by hypertonic, but not hypotonic, stimulation (3, 18). Therefore, the osmoreceptors on the cell bodies of vasopressin-containing neurons may be a different molecule from TRPV4. However, in TRPV4 KO mice, the regulatory mechanisms of vasopressin secretion are disturbed, although effects of TRPV4 on vasopressin secretion are controversial (13, 14). TRPV4 may possibly indirectly regulate vasopressin secretion through neurons expressing TRPV4. To elucidate roles for TRPV4 in the regulatory mechanisms of osmolality homeostasis, further in vivo experiments on vasopressin release are needed in WT animals.

TRPV4 in the choroid plexus (CP) is the first candidate for the action site of 4{alpha}-PDD in this study. TRPV4 in the CP is the nearest target for agonist molecules injected into the ventricle rather than other sites, such as the vascular organ of the lamina terminalis (OVLT) and the subfornical organ, where immunohistochemical studies (12–14) show the localization of TRPV4 in the CNS. The CP is known to generate the cerebrospinal fluid (CSF). Probably, TRPV4 in the CP detects osmotic pressure and regulates ion components in the CSF, resulting in control of water balance in vivo. In fact, there is some evidence that changes in the osmotic pressure/sodium concentration of the CSF regulate water intake (1, 16, 19). The organs around the anteroventral third ventricle are also important, because we found an interaction of TRPV4 and ANG II in the regulation of drinking water. All of the organs are needed for ANG II-induced dipsogenic effects after injection into the ventricle (4). Liedtke et al. (13) suggested that the OVLT is a key region for TRPV4 in osmotic regulation.

TRPV4 channels are also sensitive to heat from 30 to 42°C (7, 17). Therefore, at normal rat body temperature, the TRPV4 channel is activated, but not fully. Electrophysiological studies show that additional stimuli, for example, an agonist or osmotic stimuli, probably open TRPV4 channels synergistically, not additively (6). In in vitro studies of TRPV4-expressed cells, TRPV4 is activated by protein kinase C and tyrosine kinase (6, 15, 26). Regulation by [Ca2+]i is complex; ordinary increases activate TRPV4, but excessive increases inhibit TRPV4 (20).

In addition, our study showed an interaction between ANG II and TRPV4 pathways. It is suggested that TRPV4 is present downstream from ANG II signaling, because the TRPV4 agonist inhibited the dipsogenic effects of ANG II. Activating ANG II signaling may close TRPV4 directly or indirectly. In vitro experiments using TRPV4-transfected cells have shown that TRPV4 is closed by temperatures <30°C (7, 17), hyperosmolality (12), and excessive increases in [Ca2+]i (20). So far, there is no evidence that these mechanisms are linked to ANG II signaling. One possibility is that ANG II produces excessive increases in [Ca2+]i and then promotes protein/peptide phosphorylation, resulting in TRPV4 inactivation. In general, phosphorylation is involved in gating mechanisms of various ion channels.

Our study showed that only one-half of the ANG II-induced dipsogenic effects were inhibited by a TRPV4 agonist. We suppose that TRPV4 is not expressed on all the neurons downstream of ANG II signaling. Distribution of TRPV4 in the hypothalamus, especially along with ANG II, needs to be studied in detail. ANG II is well known to be one of the strongest dipsogenic drugs. In addition, ANG II also detects sodium concentrations in body fluids and regulates vasopressin secretion (1). Therefore, it is suggested that TRPV4 is present downstream of ANG II signaling and plays a very important role in the regulation of osmolality homeostasis.

We conclude that TRPV4 is involved in physiological regulation, via drinking behavior, of the osmotic pressure of body fluids. ANG II signaling is suggested to be present upstream of this regulation. Hyperosmolality-induced thirst is not influenced by activation of TRPV4.


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 ABSTRACT
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This study was partly supported by grant-in-aid for Scientific Research from the Ministry of Education, Culture, Sports, Science, and Technology of Japan (no. 16659060) and from Salt Science Foundation (no. 0336; Tokyo, Japan).


   FOOTNOTES
 

Address for reprint requests and other correspondence: H. Tsushima, Laboratory of Pharmacology, College of Pharmacy, Kinjo Gakuin Univ., 2-1723 Omori, Moriyama-ku, Nagoya 463-8521, Japan (e-mail: tsushima{at}kinjo-u.ac.jp)

The costs of publication of this article were defrayed in part by the payment of page charges. The article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.


   REFERENCES
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 ABSTRACT
 MATERIALS AND METHODS
 RESULTS
 DISCUSSION
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  1. Antunes-Rodrigues J, De Castro M, Elias LLK, Valenca MM, and McCann SM. Neuroendocrine control of body fluid metabolism. Physiol Rev 84: 160–208, 2004.
  2. Berl T, Siriwardana G, Ao I, Butterfield LM, and Heasley LE. Multiple mitogen-activated protein kinases are regulated by hyperosmolality in mouse IMCD cells. Am J Physiol Renal Physiol 272: F305–F311, 1997.[Abstract/Free Full Text]
  3. Bourque CW and Oliet SH. Osmoreceptors in the central nervous system. Annu Rev Physiol 59: 601–619, 1997.[CrossRef][Web of Science][Medline]
  4. Buggy J, Fisher AE, Hoffman WE, Johnson AK, and Phillips MI. Ventricular obstruction: effect on drinking induced by intracranial injection of angiotensin. Science 190: 72–74, 1975.[Abstract/Free Full Text]
  5. Clapham DE. TRP channels as cellular sensors. Nature 426: 517–524, 2003.[CrossRef][Medline]
  6. Gao X, Wu L, and O'Neil RG. Temperature-modulated diversity of TRPV4 channel gating: activation by physical stress and phorbol ester derivatives through protein kinase C-dependent and -independent pathways. J Biol Chem 278: 27129–27137, 2003.[Abstract/Free Full Text]
  7. Guler AD, Lee H, Iida T, Shimizu I, Tominaga M, and Caterina M. Heat-evoked activation of the ion channel, TRPV4. J Neurosci 22: 6408–6414, 2002.[Abstract/Free Full Text]
  8. Gunthorpe MJ, Benham CD, Randall A, and Davis JB. The diversity in the vanilloid (TRPV) receptor family of ion channels. Trends Pharmacol Sci 23: 183–190, 2002.[CrossRef][Medline]
  9. Hiyama TY, Watanabe E, Ono K, Inenaga K, Tamkun MM, Yishida S, and Noda M. Nax channel involved in CNS sodium-level sensing. Nat Neurosci 5: 511–512, 2002.[CrossRef][Web of Science][Medline]
  10. Hussy N, Deleuze C, Desarmenien MG, and Moos C. Osmotic regulation of neuronal activity: a new role for taurine and glial cells in a hypothalamic neuroendocrine structure. Prog Neurobiol 62: 113–134, 2000.[CrossRef][Web of Science][Medline]
  11. Konig LY and Klippel RA. The Rat Brain. A Stereotaxic Atlas of the Forebrain and Lower Parts of the Brain Stem. Baltimore, MD: Williams and Wilkins, 1963.
  12. Liedtke W, Choe Y, Marti-Renom MA, Bell AM, Denis CS, Sali A, Hudspeth AJ, Friedman JM, and Heller S. Vanilloid receptor-related osmotically activated channel (VR-OAC), a candidate vertebrate osmoreceptor. Cell 103: 525–535, 2000.[CrossRef][Web of Science][Medline]
  13. Liedtke W and Friedman JM. Abnormal osmotic regulation in trpv4–/– mice. Proc Natl Acad Sci USA 100: 13698–13703, 2003.[Abstract/Free Full Text]
  14. Mizuno A, Matsumoto N, Imai M, and Suzuki M. Impaired osmotic sensation in mice lacking TRPV4. Am J Physiol Cell Physiol 285: C96–C101, 2003.[Abstract/Free Full Text]
  15. Nillius B, Watanabe H, and Vriens J. The TRPV4 channel: structure-function relationship and promiscuous gating behaviour. Pflügers Arch 446: 298–303, 2003.[CrossRef][Web of Science][Medline]
  16. Osborne PG, Denton DA, and Weisinger RS. Inhibition of dehydration induced drinking in rats by reduction of CSF Na concentration. Brain Res 412: 36–42, 1987.[CrossRef][Web of Science][Medline]
  17. Patapoutian A, Peier AM, Story GM, and Viswanath V. ThermoTRP channels and beyond: Mechanisms of temperature sensation. Nat Rev Neurosci 4: 529–539, 2003.[CrossRef][Web of Science][Medline]
  18. Qiu DL, Shirasaka T, Chu CP, Watanabe S, Yu NS, Katoh T, and Kannan H. Effect of hypertonic saline on rat hypothalamic paraventricular nucleus magnocellular neurons in vitro. Neurosci Lett 355: 117–120, 2004.[CrossRef][Web of Science][Medline]
  19. Rundgren M, Denton DA, McKinley MJ, and Weisinger RS. The dipsogenic effect of intracerebroventricular infusion of hypertonic NaCl in the sheep is mainly by the Na ion. Acta Physiol Scand 127: 433–436, 1986.[Web of Science][Medline]
  20. Strotmann R, Schultz G, and Plant TD. Ca2+-dependent potentiation of the nonselective cation channel TRPV4 is mediated by a C-terminal calmodulin binding site. J Biol Chem 278: 26541–26549, 2003.[Abstract/Free Full Text]
  21. Tsushima H and Mori M. In vivo evidence that activation of tyrosine kinase is a trigger for lipopolysaccharide-induced fever in rats. Brain Res 852: 367–373, 2000.[CrossRef][Web of Science][Medline]
  22. Tsushima H and Mori M. Involvement of protein kinase C and tyrosine kinase in lipopolysaccharide-induced anorexia. Pharmacol Biochem Behav 69: 17–22, 2001.[CrossRef][Web of Science][Medline]
  23. Tsushima H, Mori M, Fujiwara N, and Moriyama A. Pharmacological characteristics of bombesin receptor mediating hypothermia in the central nervous system of rats. Brain Res 969: 88–94, 2003.[CrossRef][Web of Science][Medline]
  24. Voisin DL and Bourque CW. Integration of sodium and osmosensory signals in vasopressin neurons. Trends Neurosci 25: 199–205, 2002.[CrossRef][Web of Science][Medline]
  25. Watanabe H, Davis JB, Smart D, Jerman JC, Smith GD, Hayes P, Vriens J, Cairns W, Wissenbach U, Prenen J, Flockerzi V, Droogmans G, Benham CD, and Nilius B. Activation of TRPV4 channels (hVRL-2/mTRP12) by phorbol derivatives. J Biol Chem 277: 13569–13577, 2002.[Abstract/Free Full Text]
  26. Xu H, Zhao H, Tian W, Yoshida K, Roullet JB, and Cohen DM. Regulation of a transient receptor potential (TRP) channel by tyrosine phosphorylation: Src family kinase-dependent tyrosine phosphorylation of TRPV4 on Tyr-253 mediates its response to hypotonic stress. J Biol Chem 278: 11520–11527, 2003.[Abstract/Free Full Text]
  27. Zhang Z and Bourque CW. Osmometry in osmosensory neurons. Nat Neurosci 6: 1021–1022, 2003.[CrossRef][Web of Science][Medline]
  28. Zhang Z and Cohen DM. NaCl but not urea activates p38 and jun kinase in mIMCD3 murine inner medullary cells. Am J Physiol Renal Fluid Electrolyte Physiol 271: F1234–F1238, 1996.[Abstract/Free Full Text]



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