Fluid Transitions

Abstract

Water is critical for survival and thirst is a powerful way of ensuring that fluid levels remain in balance. Overconsumption, however, can have deleterious effects, therefore optimization requires a need to balance the drive for water with the satiation of that water drive. This review will highlight our current understanding of how thirst is both generated and quenched, with particular focus on the roles of angiotensin II, glucagon like-peptide 1, and estradiol in turning on and off the thirst drive. Our understanding of the roles these bioregulators play has benefited from modern behavioral analyses, which have improved the time resolution of intake measures, allowing for attention to the details of the patterns within a bout of intake. This has led to behavioral interpretation in ways that are helpful in understanding the many controls of water intake and has expanded our understanding beyond the dichotomy that something which increases water intake is simply a “stimulator” while something that decreases water intake is simply a “satiety” factor. Synthesizing the available information, we describe a framework in which thirst is driven directly by perturbations in fluid intake and indirectly modified by several bioregulators. This allows us to better highlight areas that are in need of additional attention to form a more comprehensive understanding of how the system transitions between states of thirst and satiety.

1. Introduction

The transition from thirsty to quenched is a common experience, but the importance of it for survival is underappreciated. Of course, we recognize that thirst and hunger are formidable motivated states. Water is critical for survival and thirst is a powerful way of ensuring that fluid levels remain in balance, much in the same way that energy is critical, and hunger is a powerful way of ensuring that sufficient nutrients are consumed to support survival. Indeed, food or water deprivation are used to create these states to drive behavior experimentally, often simply to reward a behavior or performance on a task that has nothing to do with the study of ingestive behavior. But these motivated states can also bring harm. Water or food seeking can drive risky behavior. For instance, spending too much time at an open water or food source can provide opportunities for predators. Moreover, overconsumption of food or of water can have deleterious consequences. Thus, an optimal system requires a need to balance the drive with the satiation of that drive.

2. Why do we transition to drinking?

Most research on the neural control of thirst and on the signals that stimulate drinking have been informed by the double-depletion hypothesis that was originally proposed by Fitzsimons (Fitzsimons 1973) and Epstein (Epstein 1973). According to this framework, depletion of water occurs in either the extracellular or the intracellular compartment (see Table 1 for terminology that is frequently used in the literature and in this review). When fluid is lost from the extracellular space (aka extracellular dehydration, hypovolemic dehydration, hypovolemia), it triggers mechanisms that are primarily pressure-sensing in the cardiovascular system. Key components of this detection include baroreceptors that inform the brain of the volume loss through vagal afferents, and the kidney, which detects low pressure and starts the process of synthesizing angiotensin II (Ang II; described in detail below). Because fluid lost from the extracellular compartment often carries both water and solutes in the water, a complete restoration of the lost fluid includes both thirst and sodium appetite (Daniels and Fluharty 2004, Daniels and Fluharty 2009, Santollo, Daniels et al. 2023). Unlike the loss that occurs with depletion of the extracellular space, a loss of fluid from the intracellular space (aka intracellular dehydration, osmotic dehydration) is not a true dehydration but is instead a shift of fluid from the intracellular to the extracellular space. This shift is generally caused by an increase in the osmolality of the extracellular space that draws water from the inside to the outside of the cell. Under these conditions, the physical change in shape and membrane properties of the cell, caused by the water loss, triggers a response in osmoreceptive cells that generates thirst and drives water intake. In this case, however, consumption of sodium would exacerbate the osmotic imbalance. Sodium appetite is, therefore, not present, even though it may be stimulated, but is simultaneously suppressed by oxytocin (Blackburn, Samson et al. 1993). In addition to drinking in response to immediate and rapid fluid loss, mammals are constantly losing fluids from breathing, sweating, and excretion. This is in addition to any intracellular dehydration produced by ingesting food. Animals, therefore, are constantly replacing these lost fluids in a non-emergency setting throughout the day to maintain fluid balance. We will refer to this continuous consumption, in the laboratory often measured in 24-h periods, as endogenous water intake¹.

Table 1.

Definitions and usage common in the thirst field.

Fluid State & Synonyms	Definition	Response
Intracellular Dehydration Osmotic Dehydration Intracellular Depletion	Loss of fluids from within the cell (intracellular space/compartment), generally caused by increased tonicity of the extracellular space and the resultant osmosis	Osmotic Thirst
Extracellular Dehydration Hypovolemic Dehydration Hypovolemia Extracellular Depletion	Loss of fluids and electrolytes from the plasma & interstitial fluids (extracellular compartment)	Hypovolemic Thirst Volumetric Thirst
Endogenous Intake Unstimulated Intake Need-free Intake Circadian Intake Overnight Intake Daily Intake	Intake across a period of time (often daily) that is not stimulated by an injury or experimental manipulation, but that occurs more normally

When conceptualizing the controls of fluid intake we draw inspiration from the direct and indirect controls of food intake described in Smith’s classic review on the controls of meal size (Smith 1996). Our usage of direct and indirect differs somewhat from how Smith used these terms, but the framework described here clearly borrows from Smith’s ideas. For an understanding of the controls of fluid intake, we find it helpful to define a direct control of fluid intake as one that causes the animal to transition from a state of not drinking to a state of drinking. In this framework, AngII, osmoreceptors, and baroreceptors fall into the category of direct controls. Indirect control differs from direct control in that indirect control affects the likelihood an animal will continue to drink or will stop drinking, but bioregulators that fall into the indirect category do not directly stimulate intake on their own. In Section 5, we further describe the roles of AngII, which provides direct control of fluid intake, and glucagon like-peptide 1 (GLP-1) and estradiol, both of which provide indirect control of fluid intake. In addition to directly or indirectly influencing fluid intake, there exist tertiary controls of intake that form an extended neural circuit in the control of thirst and drinking. These include signals such as dopamine (Mietlicki-Baase, Santollo et al. 2021), which is released both in response to drinking but also in response to learned cues associated with fluid intake (Fortin and Roitman 2018, Hsu, Bazzino et al. 2020). We believe that this framework helps clarify the various roles that substances can play in the control of fluid intake and provides an important mechanistic perspective for classifying any newly discovered influences on thirst and fluid intake.

3. Why do we transition to not drinking?

Although the systems that detect fluid perturbations and respond by initiating thirst have been well studied, the process by which thirst is quenched is poorly understood. Limiting intake is very important. From a clinical standpoint, excess body fluid can lead to hypertension, a risk factor for cardiovascular disease and in extreme cases, polydipsia can result in death due to hyponatremia (colloquially known as water intoxication). As mentioned above, from an ecological standpoint, spending too much time at an open water source puts an animal at risk of predation. Thus, it is important to understand how thirst is quenched and drinking bouts are terminated. An obvious and simple (although flawed) explanation of how thirst is quenched is that as the animal drinks, in response to the thirst, the water that is consumed restores the fluid that was lost, thereby removing the stimulus (the deficit) that triggered the thirst response. The flaw in this idea is clear based on studies showing that drinking, and release of vasopressin, stop well before the consumed fluid can circulate and correct the fluid imbalance. This is true in laboratory animals and in human subjects (Rolls, Wood et al. 1980, Thrasher, Nistal-Herrera et al. 1981, Arnauld and du Pont 1982, Geelen, Keil et al. 1984, Blair-West, Gibson et al. 1985, Takamata, Mack et al. 1995, Stricker and Hoffmann 2005, McKinley, Weissenborn et al. 2009). Studies using more recently developed approaches have demonstrated that some parts of the circuit respond in an anticipatory manner, with activity that corrects when the anticipated fluid intake does not occur. For instance, vasopressin-expressing cells become active when mice are not allowed to drink for a period of time, and this activity rapidly decreases when drinking begins (Mandelblat-Cerf, Kim et al. 2017), well before the water consumed would have time to reach these brain areas and decrease the osmotic imbalance caused by the water restriction. If, however, the mice are presented with an empty water vessel, the rapid decrease in activity in the vasopressin neurons is still observed, but only transiently with a similarly rapid return to the active state associated with water deprivation. Thus, the feedback from the act of drinking appears to have a learned component as well as a means to correct what is expected, and this feedback reaches the same cells that are involved in stimulating drinking. Because this is clearly more than a simple removal of the deficit that stimulated the drinking (occurring before the deficit is removed), an active process appears to be involved. How this feedback occurs is a source of current study.

Feedback from ingestion seems likely to flow through hindbrain structures. Indeed, gastric distention elicits signals that are relayed through prodynorphin (pdyn) cells in the parabrachial nucleus and the activation of these signaling cells suppresses water intake (Kim, Heo et al. 2020). It is not clear the role these cells play in overall fluid intake, however, because some reports show that the pdyn neurons are activated upon sodium deprivation and activation of these neurons increases sodium intake (Lee, Augustine et al. 2019, Gasparini, Resch et al. 2021). Other studies on gut-brain feedback related to drinking suggest that gastric osmolality is an important signal in thirst satiation (Zimmerman, Huey et al. 2019). But signals from the gut are not the only means by which intake is suppressed. Indeed, gastric bypass in rats failed to have any observed impact on fluid intake after a variety of challenges (Marshall, Santollo et al. 2014) and it has been known since the 1950s that water consumed by mouth is more satiating than water infused into the gut (Miller, Sampliner et al. 1957). The sensing of water in the mouth is detected at least in part via acid-sensing taste receptor cells (Zocchi, Wennemuth et al. 2017) and signals arising from these cells appear to be important for thirst-related forebrain activation (Augustine, Gokce et al. 2018). Selective activation of these cells alone is not, however, satiating (Zocchi, Wennemuth et al. 2017), suggesting that there is an additional oropharyngeal signal required for the satiation caused by drinking. Nevertheless, there appear to be signals for fluid intake termination that arise from various levels of the orogastric tract and it seems plausible to hypothesize that some of the same neural circuits that carry satiety signals from the gut are overlapping with those that carry signals from the mouth. This is an area that is ripe for exploration.

4. Measuring intake

A full consideration of our current understanding of the systems that govern hunger and thirst is informed by ways that scientists have studied these motivated states and the intake they drive. The amount (volume/weight) of water or food consumed is fairly simple to measure by providing a known quantity to a laboratory animal and measuring what remains after some period of time. Modern refinements have used a variety of ways to improve the time resolution of these measures (e.g., contact lickometers, beam break detection, and load cells that allow for frequent weighing of food hoppers without disturbing the animal), allowing for attention to the details of the patterns within a bout of intake. This has allowed scientists to interpret the behavior in ways that clarify the physiological controls of the behavior. Differences in eating or licking patterns, for example, have provided ways to test hypotheses about the impact of a manipulation on different properties of the substance being ingested. For example, measures of licking patterns can be used to test hypotheses about the effect of a drug on satiation. Indeed, decades of research and fine analyses of lick patterns show that a manipulation that affects the hedonic value (orosensory) of a substance will have a primary effect on the number of licks in a licking burst (Davis 1989, Davis, Smith et al. 1999). In contrast, a manipulation that impacts satiation during a bout of drinking will have a primary effect on the number of licking bursts within a bout of drinking (Davis 1989, Davis, Smith et al. 1999). Other measures, such as latency to consume food or water after the experimental manipulation, can provide additional information about the onset or decline of the drive state. These kinds of analyses have been critical in developing an understanding of the systems that govern intake, however, the utility of drinking microstructure analysis is most informative when focusing on the indirect controls of intake. This is because, in the framework presented earlier, a direct control of intake (e.g. AngII) causes the animal to transition from a state of not drinking to drinking. In that scenario both burst number and burst size will obviously differ from when the animal is not drinking because there are no licks to form bursts in the absence of drinking. These analyses are, nonetheless, very helpful in understanding the nature of indirect controls of intake and whether they have a primary effect on the hedonics or satiation.

5. Bioregulators

A variety of chemical signals form or act upon the circuits that drive thirst and thirst satiation. Some of these have been identified as controlling both eating and drinking (Taylor, Bagley et al. 2005, Samson, White et al. 2007, Mietlicki, Nowak et al. 2009, McKay, Kanoski et al. 2011, Yosten, Redlinger et al. 2012, Santollo and Daniels 2015, Mietlicki-Baase, McGrath et al. 2017, Kurt, Woodworth et al. 2019, Kurt, Kodur et al. 2022), while others are more selectively relevant for control of fluid (e.g. atrial naturietic peptide (McCann, Gutkowska et al. 1994) or food (e.g., CCK (Kraly, Carty et al. 1978)) intake. Although the bioregulators that affect thirst with or without effects on hunger, are numerous, several key bioregulators have emerged (AngII, glucagon-like peptide-1, and estradiol). This is not to say that these are the most important bioregulators, but they are arguably the best studied. We have, therefore, selected them for review here. Indeed, learning how these bioregulators control intake reveals important aspects of the system. In this respect, directionality is important to consider, but the direction of the observed response may reflect different mechanisms. For instance, upon discovering that a particular bioregulator stimulates intake, we should not jump to the conclusion that termination of intake involves removal of that bioregulator. In colloquial terms, it is important to think of the control of intake like we would the accelerator and brake functions in an automobile. The automobile can slow down by removing the accelerator or by applying the brake (or by both actions), but it would be irresponsible to observe a slowing automobile and assume that one or the other was responsible for the slowing without more information. Likewise, as we learn more about the bioregulators that are involved in the control of fluid intake, we are repeatedly finding separate roles of complicated signals in increasing and decreasing intake (Figure 1).

Figure 1.

Direct and indirect controls of thirst and fluid intake. Thirst and the resulting fluid intake are initiated by direct controls that are responsible for signaling a fluid deficit in the intracellular or extracellular space. Indirect controls modify the intake that occurs in response to the direct controls by potentiating or attenuating the impact of the direct controls.

5A. Angiotensin II

It is arguable that AngII is the most recognized and well characterized bioregulator of water intake. Stimulation of drinking by AngII has been well documented in the literature and is a key signaling peptide in response to a deficit in extracellular volume (Fitzsimons and Simons 1969, Epstein, Fitzsimons et al. 1970, Hsiao, Epstein et al. 1977). The loss of extracellular fluids is detected by both aortic and carotid baroreceptors along with juxtaglomerular and macula densa cells of the kidney. The kidney releases the enzyme renin, which cleaves the prohormone angiotensinogen to angiotensin I, which is then converted to the active hormone AngII by angiotensin converting enzyme (ACE). AngII has myriad physiological effects to restore blood volume, including activating central angiotensin type 1 receptors (AT1R) in circumventricular organs, specifically the subfornical organ (SFO) and organum vasculosum laminae terminalis (OVLT), which stimulate water intake. The brain also locally synthesizes AngII in response to AngII binding, and this centrally produced AngII then acts in a transmitter-like manner to stimulate water intake (Sigmund 2012, Coble, Grobe et al. 2015); however, whether this centrally produced AngII is necessary for the stimulation of intake remains unclear. Various transgenic models that overexpress brain renin-angiotensin system (RAS) signaling demonstrate that AngII neurotransmission increases endogenous water intake. Overexpression of SFO AngII or selective production of SFO AngII doubles daily endogenous water intake in mice, but this intake can be prevented by an AngII receptor antagonist (Sakai, Agassandian et al. 2007, Coble, Cassell et al. 2014). Furthermore, transgenic mice with brain RAS hyperactivity have increased daily endogenous water intake, which is mediated by an increase in aldosterone production. Adrenalectomy in these mice restores daily water intake to the levels observed in wild-type mice (Grobe, Grobe et al. 2010). Additional studies are necessary to understand the interactions between peripheral and central AngII signaling in controlling water intake and if central AngII signaling is necessary for drinking.

Pharmacological, optogenetic, and knockout studies all support the role of AngII in stimulating water intake. Ventricular, SFO, or OVLT injection of AngII drive water intake in a dose dependent manner in rodents (Epstein, Fitzsimons et al. 1969, Epstein, Fitzsimons et al. 1970, Fitzsimons 1971, Simpson and Routtenberg 1973, Xu and Xinghong 1999, Vento and Daniels 2014, Santollo, Torregrossa et al. 2017) and blockade of the type 1 AngII receptor (AT1R) with the antagonist losartan inhibits AngII-induced water intake (Rowland, Morien et al. 1996, Grippo, Kirby et al. 2002, Crews and Rowland 2005, Daniels and Marshall 2012), especially when forebrain AT1R is targeted (Daniels and Marshall 2012). Optogenetic and chemogenetic activation of AT1R expressing cells in the SFO and OVLT also increases water intake in sated rodents (Oka, Ye et al. 2015, Matsuda, Hiyama et al. 2017, Kinsman, Simmonds et al. 2020). This apparent role of AngII as a bioregulator that triggers drinking is experimentally supported by myriad studies showing drinking in response to AngII as well as more nuanced studies showing that AngII dose-dependently decreases the latency to drink (Anke, Van Eekelen et al. 1988). Furthermore, studies from mice lacking the angiotensinogen gene demonstrate the involvement of AngII in hypovolemic thirst. While drinking in response to intracellular dehydration is intact in angiotensinogen knockout mice, there is inhibited drinking in response to an injection of a colloid, which induces extracellular fluid loss. A corresponding decrease in Fos expression in the SFO and OVLT is observed under these conditions. Surprisingly, endogenous water intake in mice lacking angiotensinogen is approximately three times larger than in wild-type controls (McKinley, Walker et al. 2008). The increased water intake appears to be mediated by impaired kidney function resulting in the inability to concentrate urine. Interestingly, restoring brain, cardiac, or adipose, but not renal, angiotensinogen corrects the renal defect, and in brain- and cardiac-replaced mice there is a reduced daily water intake, but not to wild type levels (Lochard, Silversides et al. 2003). Angiotensinogen rescue in multiple tissues may be necessary to return endogenous water intake to wild-type levels, however, if endogenous intake requires peripheral AngII, lack of the angiotensinogen would predictably lower water intake beyond that observed in wild-type mice. Therefore, based on the available knockout models, peripheral AngII likely does not play a role in endogenous water intake in mice. This is consistent with the double-depletion framework in which AngII is primarily involved in the response to extracellular depletion (more likely emergency-related), whereas osmoreceptors underly responses to intracellular depletion (more likely caused by routine daily events).

5B. Glucagon-like peptide 1

Glucagon-like peptide-1 (GLP-1) has emerged as a candidate satiety signal for both eating and drinking. Determining which GLP-1 responsive cells are important for thirst has been confounded by the overlap between eating and drinking. Initial evidence for a role for GLP-1 in the control of drinking by pharmacological manipulation of the GLP-1 receptor (GLP-1R) was compelling, but did not rule out the possibility that the decrease in drinking was secondary to a decrease in food intake (Navarro, Rodriquez de Fonseca et al. 1996, Tang-Christensen, Larsen et al. 1996, Wang, Edwards et al. 1998, Larsen, Fledelius et al. 2001). Subsequent studies performed this test by measuring the effect of GLP-1R agonists on drinking in rats that were not allowed to eat, showing that the fluid intake suppression by GLP-1R agonist treatment persisted, even if there was no decrease in food intake (McKay, Kanoski et al. 2011). Importantly, drinking microstructure analysis demonstrates that the inhibitory effect of GLP-1 is due to changes in postingestive feedback. For example, GLP-1R agonist treatment reduces water and saline intake via reductions in burst number, with no change in burst size (McKay and Daniels 2013). Furthermore, blocking endogenous GLP-1 signaling with the antagonist exendin-9 increases water intake via an increase in burst number, again with no change in burst size (McKay, Galante et al. 2014). Analysis of overnight intake after central injection of a GLP-1R agonist found a treatment-associated difference in both burst number and burst size, but the length of the test leaves open the possibility that one preceded the other (Brakey, Schatz et al. 2023). Nevertheless, the available data provide strong licking microstructure-based evidence to support the role of GLP-1 as a satiety signal.

Additional experiments provide evidence for clear differences between the effect of GLP-1 on eating and on drinking. For instance, levels of circulating GLP-1 do not increase after water-deprived rats are allowed to drink, but do increase after eating in food-deprived rats (McKay, Galante et al. 2014). Measures of brain GLP-1 expression were, however, affected by either eating or drinking (McKay, Galante et al. 2014). Other findings indirectly support the idea that eating involves both central and peripheral GLP-1 whereas drinking more selectively involves the central GLP-1 system. For instance, maintenance on a high-fat diet attenuates the food intake suppression by GLP-1R agonist treatment, but only when the drug is delivered centrally (Williams, Hyvarinen et al. 2011). In contrast, maintenance on high-fat diet had no effect on the water intake suppression by a GLP-1R agonist (Volcko, Carroll et al. 2020). Whether or not the central GLP-1 system has elements that are selective for food or for water intake control is in need of additional study.

More recently the Brattleboro rat, a rodent model of diabetes insipidus, has emerged as a potential way to tease apart GLP-1 circuits involved in eating and drinking. Brattleboro rats are severely polydipsic because of marked polyuria. The high level of drinking occurs without any differences in the size of licking bursts, but is entirely because Brattleboro rats have more bursts of licking throughout the day (Brakey, Schatz et al. 2023), suggesting that Brattleboro rats lack the satiating feedback from drinking that occurs in wildtype rats. This difference in satiety, and previous work suggesting that GLP-1 is important for intake satiation, support the hypothesis that GLP-1 function is different in Brattleboro rats. Indeed, Brattleboro rats are hypersensitive to the fluid intake suppression caused by treatment with a GLP-1R agonist (Brakey, Schatz et al. 2023). This alone makes them an interesting model for the study of GLP-1, but the finding that they are comparable to wildtype rats in the suppression of food intake after GLP-1R treatment opens the exciting possibility that whatever aspect of GLP-1 that is different in Brattleboro rats will reveal drinking-specific parts of the GLP-1 system. Additional research using this model is underway.

5C. Estradiol

Historically, the role of estrogens in controlling water intake was thought to be inhibitory, like its well characterized anorexigenic effect. Across the estrous cycle, endogenous 24-h water intake is reduced on the day of estrus (Antunes Rodrigues and Covian 1963, Tarttelin and Gorski 1971, Findlay, Fitzsimons et al. 1979, Danielsen and Buggy 1980, Eckel, Houpt et al. 2000, Santollo, Edwards et al. 2021). Ovariectomy increases endogenous 24-h water intake (Tarttelin and Gorski 1971, Santollo, Edwards et al. 2021), and estradiol (the most potent circulating estrogen) replacement decreases intake (Spiteri, Drewett et al. 1980, Santollo, Edwards et al. 2021). Because rats are prandial drinkers, it is important to ask if the effect on fluid intake is direct or if it occurs secondary to a decrease in food intake. Two key findings answer this question. First, in rats, the estrus-related decrease in 24-h water persists when rats are not allowed to eat (Santollo, Edwards et al. 2021). Second, in ovariectomized (OVX) guinea pigs, restricting food intake to 30% less than ad libitum levels abolished the anorexigenic effect of exogenous estradiol treatment but the anti-dipsogenic effect remained intact (Czaja, Butera et al. 1983). Thus, even when food and water intakes are uncoupled, the effect of estrogens on water intake remains. Drinking in response to dehydration is also inhibited by estrogens. Although estradiol does not appear to be involved in osmotic thirst (Findlay, Fitzsimons et al. 1979, Jonklaas and Buggy 1984, Krause, Curtis et al. 2003, Mecawi, Lepletier et al. 2008), multiple reports demonstrate estradiol’s impact on hypovolemic thirst. Multiple laboratories have reported that estradiol decreases water intake in response to 24-h water deprivation in ovariectomized rats (Krause, Curtis et al. 2003, Mecawi, Lepletier et al. 2008, Vilhena-Franco, Mecawi et al. 2016, Howell, Edwards et al. 2024). Although water deprivation clearly causes dehydration, estradiol also inhibits water intake in response to treatments that model portions of dehydration and its relevant signals. For instance, AngII is a powerful dipsogen and part of the response to extracellular dehydration. Although drinking occurs after AngII injection throughout the estrous cycle, drinking after AngII is lowest on the day of estrus than it is on other days of the estrous cycle (Findlay, Fitzsimons et al. 1979, Danielsen and Buggy 1980). Furthermore, in OVX rats, estradiol treatment reduces AngII-induced drinking, demonstrating that estradiol and not other ovarian hormones mediates the change in intake (Fregly 1978, Fregly and Thrasher 1978, Jonklaas and Buggy 1984, Kisley, Sakai et al. 1999, Santollo and Daniels 2015, Santollo, Marshall et al. 2016, Santollo, Collett et al. 2021). Estradiol also reduces water intake that is caused by other treatments that stimulate the renin-angiotensin system. For example, estradiol decreases drinking in OVX rats treated with isoproterenol (Thrasher and Fregly 1978, Krause, Curtis et al. 2003, Graves, Hayes et al. 2011), renin (Thrasher and Fregly 1977), furosemide + captopril (Mecawi, Lepletier et al. 2008), or 24-h sodium deprivation (Mecawi, Lepletier et al. 2008), each of which stimulates the release of endogenous AngII. The drinking effects of estradiol occur through activation of estrogen receptor alpha (Santollo, Marshall et al. 2016), similar to its anorexigenic effect (Santollo, Wiley et al. 2007, Thammacharoen, Geary et al. 2009), via both nuclear and membrane-associated receptors (Santollo, Marshall et al. 2013). Of note, the drinking stimulated by these various dipsogens occurs in the absence of food intake, providing additional support for a primary inhibitory role of estradiol in the controls of water intake.

The estrogenic inhibition of water intake is consistent with the general sense that estrogens promote satiety or otherwise serve as a satiety signal. Indeed, this notion is supported by findings related to drinking microstructure patterns. Both estradiol and the ERα agonist PPT reduce endogenous 24-h water intake in OVX rats via reductions in both burst number and burst size (Santollo, Marshall et al. 2013, Santollo, Marshall et al. 2016). As previously discussed, changes in burst number are associated with post-ingestive feedback signals, implicating estradiol in reducing water intake via increasing satiety-specific or satiety-related (i.e. gastric stretch) signals. In addition, estradiol and PPT-treatment in OVX rats reduce AngII-stimulated water intake through changes in burst number, providing additional support for the notion that estradiol increases satiety or satiety-related signaling to enhance water intake termination. The critical satiety signals that estradiol augments are unknown, which may not be surprising given that the critical satiety signals for thirst are overall poorly understood. Past research focused heavily on the hypothesis that direct interactions between estradiol and AngII were responsible for the attenuated drinking responses. These efforts, however, failed to find the hypothesized interactions. For example, estradiol does not influence plasma renin activity in response to isoproterenol treatment (Krause, Curtis et al. 2006), suggesting estradiol does not influence endogenous AngII production. Estradiol also fails to influence AT1R expression when changes in body weight are considered (Krause, Curtis et al. 2006, Santollo, Marshall et al. 2016). Although studying estradiol-AngII interactions was a reasonable approach, it is now unsurprising that estradiol-AngII interactions do not underlie the estrogenic inhibition of water intake because of the newer appreciation that estradiol is more related to satiation and AngII is more involved in stimulation of water intake. That does not mean that estradiol acts alone. Indeed, recent data suggest that estradiol enhances the sensitivity of the anti-dipsogenic effect of GLP-1 (Howell, Edwards et al. 2024). This is a promising step in identifying the bioregulators that estradiol augments and presents an important opportunity for future research investment.

Recent data from our group has drastically expanded our understanding of the estrogenic control of water intake. Despite decades of research describing the estrogenic inhibition of water intake, in 2021 we demonstrated that under certain testing conditions, estradiol can increase water intake in OVX rats (Santollo, Edwards et al. 2021). In the absence of food, estradiol increases endogenous 24-h water intake, through activation of ERβ (Santollo, Edwards et al. 2021, Santollo and Edwards 2023). Exogenous estradiol also increases rebound drinking, in response to an acute pharmacological suppression of intake, in OVX rats (Santollo, Edwards et al. 2021). In both of these experiments, the drive to drink was experimentally reduced. In the first experiment, removal of food caused an approximate 60% reduction in 24 h water intake and in the second experiment our drug manipulation abolished water intake for a minimum of 4 hours. This has led us to hypothesize, that in contexts where the motivation to drink is low, or suppressed, E2 acts to enhance water intake. Although not measured, the differences in intake that were observed, may be a function of latency to drink. In support of this idea, experiments using OVX rats found that estradiol reduces the latency to drink in response to hypertonic saline treatment, despite no hormone-mediated change in total volumetric consumption (Jones and Curtis 2009). More work is needed to explore the role estradiol may have in changing drinking latency. Consistent with estradiol’s inhibitory role in water intake, the increase in endogenous 24-h water intake is also associated with a change in burst number. This change, of course, is in the opposite direction of the anti-dipsogenic effect, suggesting that estradiol is weakening the satiety signals that would normally terminate intake. Whether or not estradiol augments the same satiety signals, just in different directions, to both increase and decrease water intake or if different satiety signals are involved in increasing vs decreasing water intake, is an open question.

6. Circadian and Seasonal Control of Water Intake

A discussion of transitions between hydration and dehydration need also to consider circadian and seasonal influences. Nocturnal rodents consume most of their water intake during the dark (active phase), similar to their patterns of food intake; however, the nocturnal nature of water intake is not strictly the result of prandial drinking. When food access is restricted to the light phase, the majority of water intake still occurs during the dark phase (Spiteri 1982). In support for the circadian control of water intake, lesions of the suprachiasmatic nucleus (SCN), the master circadian clock, result in an equal distribution of water intake across the active and inactive phase in Siberian Hamsters (Bittman, Bartness et al. 1991). But how the clock controls water intake is unclear and an open area of research. Interestingly, it has been noted that both rats and mice show an increase in water intake just before the inactive phase (Spiteri 1982, Johnson, Beltz et al. 2003, Gizowski, Zaelzer et al. 2016). In an eloquent series of experiments, Bourque and colleagues demonstrated that this anticipatory water intake is mediated by the central clock SCN neurons and not physiological need. This anticipatory water intake maintains hydromineral balance throughout the inactive phase (Gizowski, Zaelzer et al. 2016). Specifically, SCN vasopressin neurons excite OVLT neurons via V1a receptors during the 2 h period prior to lights on, which drives water intake. Expanding upon the 24-h control of water intake, seasonal changes in water influx have been reported in a variety of desert dwelling rodents (Stallone 1979, Degen 1991, Nagy 1994). These changes, however, are secondary to changes in dietary content. In these environments, the winter vegetation has a significantly greater water content resulting in greater water influx in the winter, compared to summer months. Whether seasonal cycles influence water intake independent of the water content of the diet in animals living outside of desert climates is unclear and an interesting area of future research.

7. Concepts and future thoughts

Although significant progress has been made in our understanding of the control of thirst and thirst satiation, there are some seemingly important unanswered questions. Despite decades of research on the role of AngII in drinking, it is still not clear if centrally generated AngII is necessary for drinking and if so, is it specific to propagating the signal generated by circulating AngII acting on the brain? It seems likely that central AngII is important, given its position in the circuit and the clear importance of AngII in general, but a specific role has yet to be demonstrated.

The demonstration of numerous bioregulators in the control of thirst is clear progress in our understanding of how thirst is controlled. With the knowledge that these bioregulators play some role in thirst, we now must try to better understand which of the dipsogenic indirect controls act by stimulating drinking or by removing an inhibition of drinking. Conversely, we need to determine if substances with anti-dipsogenic properties act by inhibiting stimulation or by terminating drinking (Figure 1). Studies of estradiol could be especially promising in this respect because it either increases or decreases drinking depending on the context.

Layered upon the complexity of understanding the transition from thirsty to quenched, we must also recognize the intertwined role that food intake plays. In addition to being another drive state that transitions from on to off (or off to on), the frequent co-occurrence of hunger and thirst and the overlapping circuits and bioregulators involved in both create additional complications in studying each in isolation. This is clear in the attempts to understand the roles of estrogens and of GLP-1 in the control of intake. Recent work with the Brattleboro rat suggests one approach to disentangling these drive states, at least with respect to GLP-1, but this approach lacks the ease of genetic manipulation that is abundant in studies using mice, and these types of approaches in rats are still well behind those that are common in mice. A mouse model of the Brattleboro rat could be useful, but to the best of our knowledge, attempts to generate that model have largely been unsuccessful and we are unaware of any tests of GLP-1 effects in other models of diabetes insipidus.

From a broader perspective, the study of drinking has been a critical part of how we understand motivated behaviors. Studies of drinking have helped form ideas about consummatory vs appetitive phases of behavior and have provided a useful and reliable model for the study of peptide and traditional transmitters in the control of behavior. Although we have a strong understanding of the stimuli that trigger drinking, our understanding of the termination of drinking remains somewhat murkier. A better understanding of this, along with a better understanding of how food intake satiation occurs, will allow for a potentially important comparison that will likely reveal overlapping and non-overlapping substrates. This is critical for a full understanding of how the brain functions. We are also in a state of imbalance in our understanding of how experience shapes eating and drinking, with much more attention being paid to the impact of experience on food choice and reactions to energy imbalance. This may be largely because drinking has been considered more innate and less flexible. Studies of sensitization and desensitization of both water and sodium intake (Pereira, Menani et al. 2010, Vento and Daniels 2010, Vento and Daniels 2012, Vento, Myers et al. 2012, Hurley and Johnson 2013, Hurley, Thunhorst et al. 2014, Hurley, Zhang et al. 2014, Santollo, Whalen et al. 2014, Vento and Daniels 2014, Pereira-Derderian, Vendramini et al. 2016, Postolache, Santollo et al. 2017) show that this is perhaps an unfair characterization. Likewise, early life experience shapes drinking behavior. This is clear from studies of preweanling rats (Myers, Arnold et al. 1997) and from rats only allowed to drink saline or sucrose solutions from weaning until adulthood (Volcko, Brakey et al. 2020). But these studies barely scratch the surface of how learning and experience may shape water intake. Accordingly, the role of learning is ripe for study. Related to this, understanding differences in reward processes that drive water and that drive food intake could be productive avenues to a better understanding of both intake control and reward. Food intake appears related to both positive and negative reinforcement. Water, however, seems to be more related to negative reinforcement. In more colloquial terms, we often eat for pleasure even when we are not eating to ease the pang of hunger, but deriving pleasure from water intake seems far more limited to times when that water is quenching a thirst. Of course, this may be an apples to oranges (or chocolate to dry toast) comparison. The foods most often consumed as positive reinforcers are a select subset of those eaten for negative reinforcement, but water is always just water. Without more work in this area, the answers to these questions and a better understanding of the system will remain a mystery.

Highlights.

• Thirst and water intake are under direct and indirect controls

• Stimulation of thirst is better understood than quenching thirst

• Numerous bioregulators influence thirst and water intake

• Drinking microstructure analysis helps understand how bioregulators change intake