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¹ Department of Anatomy and Neurobiology, Washington University School of Medicine, St Louis, MO 63110, USA

	Abstract

Top Abstract Introduction Why do we need... What is sodium appetite... What causes sodium appetite? Where is sodium appetite... Summary Future directions References

 
Sodium appetite, the behavioural drive to ingest salt, is stimulated by prolonged physiological sodium deficiency in many animal species. The same neural mechanisms that are responsible for sodium appetite in laboratory animals may influence human behaviour as well, with particular relevance to the dietary salt intake of patients with diseases such as heart failure, renal failure, liver failure and salt-sensitive hypertension. Since the original experimental work of Curt Richter in the 1930s, much has been learned about the regulation of salt-ingestive behaviour. Here, we review data from physiology, pharmacology, neuroanatomy and neurobehavioural investigations into the stimulatory and inhibitory signals that regulate sodium appetite. A rudimentary framework is proposed for the brain circuits that integrate peripheral information representing the need for sodium with neural signals for the gustatory detection of salt in order to drive a motivated ingestive response. Based on this model, areas of remaining uncertainty are highlighted where future information would allow a more detailed understanding of the neural circuitry responsible for sodium appetite.

(Received 14 August 2007; accepted after revision 26 October 2007; first published online 2 November 2007)
Corresponding author J. C. Geerling: Washington University School of Medicine Department of Anatomy & Neurobiology, Box 8108, 660 South Euclid Avenue, St Louis, MO 63110, USA. Email: geerlinj{at}msnotes.wustl.edu

	Introduction

Top Abstract Introduction Why do we need... What is sodium appetite... What causes sodium appetite? Where is sodium appetite... Summary Future directions References

 
‘Some seek not gold, but there lives not a man who does not need salt.’ Cassiodorus

Sodium chloride, more commonly known as salt, is a necessary part of our diet, yet sodium is more than just an essential micronutrient. Wars have been fought over the control of environmental sources of salt, which was a more valuable commodity than gold at various points in history (Kare et al. 1980). The English word ‘salary’ derives from the Latin word for salt (salarium), which was once used to pay Roman soldiers. Today, a productive person is said to be ‘worth his salt’.

Other prominent references to salt are sprinkled throughout history. The poet Homer referred to it as the ‘divine substance’, and Plato noted that salt is ‘especially dear to the gods’ (Kurlansky, 2002). The word ‘salt’ appears more than 50 times in the Bible, in well-known verses such as ‘You are the salt of the earth’ (Matthew 5: 13). Shakespeare's King Lear was based upon a popular European folk story in which a king banishes his daughter for saying that she loves him ‘like salt’, but then forgives her after attempting to live without it.

Salt remains a major ingredient (some would argue that it is too major an ingredient) in our modern diet. We do not crave other minerals, such as magnesium, iodine or potassium, in the way that we desire the taste of sodium. The controversial link between dietary sodium and hypertension (elevated blood pressure) has led a number of groups, including the American Medical Association, to call for sweeping reductions in dietary salt content, particularly in processed foods. While epidemiologists clash over the harms and benefits of different amounts of dietary sodium in healthy versus hypertensive individuals (Taubes, 1998; Mitka, 2004), this review will consider a separate question: why is salt so appealing? Here, we review many of the physiological conditions and neural mechanisms discovered in laboratory animals to help explain the allure of salt.

	Why do we need to eat salt?

Top Abstract Introduction Why do we need... What is sodium appetite... What causes sodium appetite? Where is sodium appetite... Summary Future directions References

 
Body fluid balance is meticulously regulated by neuroendocrine control systems. After a change in the volume or content of the extracellular fluid (ECF; which includes blood plasma), these control systems enact appropriate compensations to bring it back within narrow limits. In the equations of body fluid balance, the two primary variables are water and sodium. Balance is maintained through complementary adjustments in their ingestion and excretion.

The control systems that regulate the intake and excretion of water are the primary means for adjusting the concentration of solutes in the extracellular fluid, whereas control of both water and sodium is necessary for maintaining a volume of blood sufficient for optimal tissue perfusion by the heart (Verbalis, 2003). The basic regulatory mechanisms for controlling water intake (thirst), water excretion and sodium excretion are fairly well characterized and widely appreciated (Andersson, 1978; Stricker & Sved, 2002; McKinley et al. 2004), but less is understood about the regulation of sodium intake.

Sodium is, by far, the most abundant extracellular solute. The osmolarity of the extracellular fluid (including blood plasma) is dictated primarily by the concentration of sodium and its attendant anions (Verbalis, 2003). Extracellular osmolarity is meticulously maintained near a set point of roughly 290 mosmol l^–1 (the vast majority of which is composed of 140 mM Na⁺, plus its attendant anions, primarily chloride).

In order to appreciate the role of sodium in body fluid homeostasis, it is critical to understand that extracellular osmolarity is regulated primarily by the ingestion and excretion of water, whereas the volume of extracellular fluid is directly proportional to the total body content of sodium (Verbalis, 2003). That is, the total volume of extracellular fluid in the body depends largely upon the amount of sodium present in the extracellular space, around which water input and output are tailored to tightly control osmotic pressure. This regulatory arrangement is the reason that sodium must be excreted to reduce plasma volume, and it must be ingested and retained to increase plasma volume.

Expansion of the ECF volume (a requirement for growth and for replacing fluid losses) is therefore absolutely limited by dietary sodium intake. Water intake alone is adequate to replace volume losses only when the sodium concentration is elevated owing to a loss of water greater than the loss of sodium. In contrast, as shown in Fig. 1, fluid losses involving large amounts of sodium (prolonged sweating or bleeding, for example) cannot be adequately replaced by water intake alone (Stricker & Jalowiec, 1970; Nose et al. 1988).

View larger version (39K): [in this window] [in a new window]   Figure 1.  Intracellular and extracellular dehydration A, after body water is lost, the extracellular fluid (ECF) volume decreases, its solute concentration (osmolarity) increases, and cells shrink because water moves out into the now-hypertonic extracellular space, a condition referred to as ‘intracellular dehydration.’ This situation stimulates antidiuresis and thirst, and the resulting ingestion and retention of water can restore ECF volume and osmolarity to normal (note that for simplicity, this figure does not include the renal natriuresis that can occur during more prolonged dehydration, which does not restore ECF osmolarity, but does result in a minor additional loss of ECF volume, providing a minor stimulus for sodium appetite in addition to thirst). B, in contrast, ‘extracellular dehydration’ is a more significant reduction in ECF volume (with or without a change in osmolarity) due to a loss of sodium-containing fluids. In this case, the ingestion of water alone is not adequate to restore the ECF to normal. In fact, excess water ingestion will cause a decrease in ECF osmolarity (hyponatraemia), which inhibits additional intake and retention of fluids until an adequate amount of solute (sodium) is restored.

Since sodium represents less than 1% of the extracellular fluid by weight, the mass of sodium required for volume restoration is quite small relative to the amount of water that must be consumed. Accordingly, antidiuresis and thirst are the primary homeostatic drives stimulated in response to fluid loss and, initially, the most critical aspect of sodium regulation is retention by the kidney.

When the ECF volume expands, the blood pressure rises, and the increased perfusion pressure causes the kidneys to excrete more sodium (Guyton, 1991). Conversely, when the ECF volume is reduced, sodium is retained. This retention is primarily mediated by the steroid hormone aldosterone. Elevated levels of aldosterone, which is produced in the adrenal glands, can stimulate near-total conservation of sodium from the urine. This remarkable regulatory mechanism is critical for survival. Removal or gross dysfunction of the adrenal glands is invariably lethal without either exogenous replacement of aldosterone or continuous dietary supplementation of sodium (Richter, 1936; Wilkins & Richter, 1940).

Eventually, however, sodium conservation is only half the battle; the kidneys can retain only what is already present in the body. The other important aspect of this control system is the regulation of salt intake. Under normal conditions, obligatory sodium losses are small and, when necessary, the kidneys can maintain near-total urinary sodium conservation for extended periods of time. This allows animals to survive for many weeks on a sodium-free diet (Orent-Keiles et al. 1937; Fine et al. 1987a). Ultimately, however, ECF volume can be neither increased nor restored without the consumption of sodium. The chronic volume deficit and the secondary increase in plasma potassium that result from sodium deprivation lead insidiously to severe health consequences. Chronic sodium deprivation causes growth retardation, reproductive deficits, reduced muscle mass, alterations in bone composition and various other pathologies, which are eventually lethal (Orent-Keiles et al. 1937; Bursey & Watson, 1983; Fine et al. 1987a).

Put simply, normal growth requires the ingestion and retention of sodium. Without dietary salt, growth slows, reproduction fails, and animals die prematurely (Orent-Keiles et al. 1937; Fine et al. 1987a,b). In humans sustained on sodium-free nutrient infusions, bone mineralization ceases and growth stops in all tissues except fat (Rudman et al. 1975). Even short-term sodium deficiency in humans causes severe muscle cramps, loss of appetite, nausea, fatigue and considerable weight loss (McCance, 1936).

Given these severe health consequences, particularly the deficits in growth and reproduction, it should come as no surprise that a hard-wired behavioural mechanism has evolved to promote salt intake in response to a prolonged sodium deficiency.

	What is sodium appetite and why is it important?

Top Abstract Introduction Why do we need... What is sodium appetite... What causes sodium appetite? Where is sodium appetite... Summary Future directions References

 
Sodium appetite (also known as salt appetite) is a motivated behavioural state that arises in a number of species specifically as a response to sodium deficiency. As the name indicates, it drives an animal to seek and ingest foods and fluids that contain sodium. Sodium appetite is a hard-wired regulatory mechanism and, like thirst, it is vital for restoring extracellular fluid.

It is important to note that when salty foods or fluids are freely available, animals (including humans) spontaneously exhibit a baseline or ‘need-free’ level of intake in excess of any immediate need or growth requirement. This baseline ingestion of salt (and water) is more than adequate for maintaining fluid balance in the absence of significant fluid loss, and any excess sodium or water is simply excreted in the urine. The magnitude of ‘need-free’ salt intake can be influenced by prior episodes of sodium deficiency (Sakai et al. 1989), especially prenatal experience with maternal illness during pregnancy (Nicolaidis et al. 1990; Crystal & Bernstein, 1995, 1998). It remains unclear, however, to what extent spontaneous salt-ingestive behaviours engage the same brain circuits that are responsible for sodium appetite, which is operationally defined as a specific response to sodium deficiency.

Abundant anecdotal evidence for sodium appetite existed for centuries (Kare et al. 1980; Denton, 1982), but a direct experimental demonstration awaited the seminal work of Curt Richter (1936). At this time, it was recognized that removal of the adrenal glands rendered animals unable to conserve urinary sodium owing to the loss of a vital ‘mineralocorticoid’ hormone (aldosterone) produced in the adrenal cortex. Unless their diet was continually supplemented with sodium, adrenalectomized animals deteriorated rapidly and died after roughly 1 week. Richter wanted to know whether animals possess an innate behavioural mechanism that would compel them to seek and ingest extra salt if it suddenly became necessary for their survival. When he gave adrenalectomized rats access to saline, they drank greatly increased amounts, even at a high concentration (3% NaCl; roughly the concentration of seawater), which they had only sampled in small amounts prior to surgery. This dramatic behavioural change is evident in one of his original charts, reproduced in Fig. 2. The voluntary increase in salt intake by these rats was more than sufficient to compensate for their urinary sodium losses, allowing their continued survival.

View larger version (31K): [in this window] [in a new window]   Figure 2.  Curt Richter provided the first experimental evidence for sodium appetite, showing that rats greatly increase their ingestion of saline in response to a physiological deficit In these experiments, a large and continual shortage of sodium was produced by removing the adrenal glands, which produce a steroid hormone critical for sodium retention by the kidneys, aldosterone. Most compelling are the data in the right panel, showing the ingestive behaviour of a representative rat that began drinking saline in large volumes after adrenalectomy, even at a saline concentration which rats normally ingest in very small amounts, if at all (3% NaCl is roughly the concentration of seawater). These graphs were reproduced from Fig. 2 of Richter's classic paper (Richter, 1936), with permission.

Subsequent investigators identified a number of other experimental methods that produce a sustained sodium deficit (hypovolaemia) to stimulate sodium appetite without removing the adrenal glands. These methods include chronic dietary sodium deprivation (Nachman & Pfaffmann, 1963; Wagman, 1963; Contreras & Hatton, 1975; Stricker et al. 1991), peritoneal dialysis (Falk & Lipton, 1967; Toth et al. 1987), colloid-induced hypovolaemia (Stricker & Jalowiec, 1970; Stricker, 1981), and furosemide diuresis combined with short-term dietary sodium deprivation (Jalowiec, 1974; Wolf, 1982).

Appropriately, the appetite stimulated by sodium deficiency is highly specific for the taste of sodium salts (Richter & Eckert, 1938). Sodium-deficient rats consistently choose sodium over non-sodium salts (potassium, calcium, etc.), and the paired anion (chloride, acetate, etc.) has little or no effect on this preference (Nachman, 1962).

Sodium appetite is a highly motivated behavioural state. Sodium-deprived rats will perform increased amounts of work (bar pressing) for a salty reward (Wagman, 1963; Quartermain et al. 1967; McCutcheon & Levy, 1972). They will also sprint significantly faster down a runway leading to a tube of saline when they are salt-hungry (Zhang et al. 1984; Schulkin et al. 1985). Interestingly, the hedonic values of other, normally rewarding stimuli, such as sugar, appear to decrease in concert with the increasing appeal of sodium (McCance, 1936; Grippo et al. 2006; Morris et al. 2006). In fact, the normal preference for sugar over salt reverses during sodium deficiency, such that rats will ingest more saline than glucose or other sugary solutions (Smith et al. 1968; Nozaki et al. 2002). When given the choice, sodium-deprived rats will even choose the taste of salt over moderate intensities of directly rewarding brain stimulation (Conover et al. 1994).

Although the potency of sodium appetite varies widely across species, it is a highly conserved behavioural response. Once dismissed as a phenomenon unique to select herbivores exhibiting salt-seeking behaviours that are readily observed in the wild (Kare et al. 1980), definitive evidence for sodium appetite has been found in a wide variety of species, including mice (Denton et al. 1990), rats (Richter, 1936), rabbits (Denton et al. 1985), pigeons (Epstein & Massi, 1987), kangaroos (Blair-West et al. 1968), sheep (Denton & Sabine, 1961), goats (Baldwin, 1969), cattle (Bell & Sly, 1979; Blair-West et al. 1997), horses (Houpt et al. 1991), monkeys (Schulkin et al. 1984; Denton et al. 1993) and humans (Wilkins & Richter, 1940; Takamata et al. 1994; Cruz et al. 2001; Kochli et al. 2005).

Locating salt remains a necessity for many animals living in the wild, particularly herbivores in sodium-impoverished environments (Blair-West et al. 1968), but not for humans. Sodium is now widely available and present in copious amounts in our diet, such that most people never experience a deficit that is prolonged or severe enough to stimulate sodium appetite. Older clinical trials that failed to produce hypovolaemia (a sustained sodium deficiency) have been cited as evidence against the existence of sodium appetite in humans (Bertino et al. 1982; Beauchamp et al. 1983, 1987), but clinical observations and subsequent experiments involving true sodium deficiency suggest otherwise.

In 1940, Wilkins and Richter described the salt cravings of a child suffering from undiagnosed adrenal disease (probably the severe salt-wasting form of congenital adrenal hyperplasia; see Kochli et al. 2005). As described in this fascinating case report, the boy expressed an extreme and persistent desire for both salt and water from a very early age (Wilkins & Richter, 1940). Even before he could speak, he would demand salt on everything he ate, and ‘salt’ was one of the first words he learned. He did not care for sugar or sweets, preferring instead to eat pure salt or salty foods. Like Richter's adrenalectomized rats, this boy needed to ingest sodium regularly just to survive; shortly after he was forced to eat a standard hospital diet and denied access to salt, he died (his adrenal disease was not diagnosed until autopsy).

More recent investigators have provided evidence for increased voluntary salt intake by children with salt-wasting congenital adrenal hyperplasia, in which adrenal steroid production is lacking as a result of mutations in the gene encoding the enzyme 21β-hydroxylase (Kochli et al. 2005). Similarly, children with Gitelman's syndrome, a genetic salt-wasting disorder caused by a defective renal sodium transporter, eat increased amounts of salt (relative to their unaffected family members) in order to maintain an ECF volume adequate to maintain a low-normal blood pressure (Cruz et al. 2001).

Increased sodium appetite has also been reported in adults: in renal patients after dialysis (Leshem & Rudoy, 1997) and in normal subjects tested after sustained sodium deficiency (Takamata et al. 1994). In the latter study, healthy adult subjects were sodium-depleted by exercise in a heated room, which caused large sweat losses of sodium. After remaining sodium deficient over the following day (with only water and salt-free food available for consumption), subjects' palatability ratings for the taste of concentrated saline peaked with a time course resembling the delayed appearance of sodium appetite in laboratory animals after peritoneal dialysis or drug-induced diuresis (Ferreyra & Chiaraviglio, 1977; Rowland & Morian, 1999). More rigorous laboratory studies of sodium appetite, in humans experiencing a more prolonged sodium deficit, would be useful for determining the influence of sodium appetite on human salt consumption in various physiological and pathophysiological conditions.

Understanding the mechanisms that influence salt intake in humans is important because excess sodium has damaging effects in select groups of people. In most healthy individuals, blood pressure only varies slightly (if at all) even with large changes in dietary salt intake (Graudal et al. 1998; Taubes, 1998; Sacks et al. 2001). Nonetheless, excess dietary sodium is an important risk factor for increasing numbers of patients who suffer from a variety of chronic medical conditions. Salt restriction is a cornerstone of successful therapy for many diseases, including congestive heart failure, liver failure, kidney failure and salt-sensitive hypertension. Unfortunately, patients with these diseases are notoriously non-compliant when they are told to eat a low-sodium diet (Korhonen et al. 1999; Ohta et al. 2004). The normal palatability of sodium certainly contributes to their non-compliance, but it is also likely that many patients experience paradoxical increases in sodium appetite as an attendant symptom of their disease (Langford et al. 1977; DiNicolantonio et al. 1982; Hurley et al. 1987; Leshem & Rudoy, 1997; Francis et al. 2001).

When the mechanisms responsible for sodium appetite are better understood, it may be possible to design evidence-based therapies that aid patients in reducing their hunger for salt. Some currently available drugs are already known to decrease sodium appetite in animal models. For example, the mineralocorticoid receptor antagonist spironolactone, whose beneficial effects in the treatment of human heart failure are firmly established (Pitt et al. 1999), also prevents the increased salt intake normally exhibited by rats in heart failure (Francis et al. 2001). Patients may live longer, healthier lives if this or other inhibitors of sodium appetite can assist them in reducing their salt intake, particularly with drugs already shown to decrease morbidity and mortality (Pitt et al. 1999, 2003).

Beyond any therapeutic potential, however, understanding the neural foundations of sodium appetite will expand our understanding of appetitive brain circuits in general. This most basic of appetites is an important model for other motivated behaviours. The brain circuits that detect the need for sodium and drive salt intake may share components in common (or in parallel) with circuits that control other appetitive drives, such as hunger and thirst. A full understanding of sodium appetite will require an integrated appreciation of the physiological, endocrinological and neural changes that stimulate it and, ultimately, identification of the specific brain circuits that integrate these inputs and drive the consumption of salt.

	What causes sodium appetite?

Top Abstract Introduction Why do we need... What is sodium appetite... What causes sodium appetite? Where is sodium appetite... Summary Future directions References

 
Sodium appetite characteristically increases after a prolonged period of sodium deficiency, but the exact mechanisms responsible for this behavioural change remain incompletely understood. As explained above, a reduction in body sodium content is equivalent to a loss of extracellular fluid volume. Within 1–2 h after the onset of hypovolaemia, an increase in thirst becomes evident as water is consumed in direct proportion to the volume of fluid lost from the extracellular space (Fitzsimons, 1961; Stricker, 1968). In contrast to thirst, the appetite for sodium does not increase until well after the onset of hypovolaemia; many hours or days later, depending upon the experimental model (Fig. 3). Initially, this distinction led to the ‘reservoir hypothesis’ suggesting that, analogous to the central osmoreceptors cells that shrink as a result of increased ECF osmolarity and stimulate thirst, there exist specialized cells or tissues that produce a delayed stimulus for sodium appetite as they gradually release extra sodium into the ECF down an increasing concentration gradient (Wolf & Stricker, 1967).

View larger version (11K): [in this window] [in a new window]   Figure 3.  In response to a physiological sodium deficit, sodium appetite increases in a delayed fashion An increased drive to ingest salt does not become evident until hours or days after a more pronounced increase in thirst. The time scales on this graph are arbitrary; the potentiation of salt intake can increase over a period of days, as occurs in response to dietary sodium deprivation, or over as little as 12–48 h, as occurs after acute hypovolaemia due to blood loss, diuresis, dialysis, or other means.

Another difference between sodium appetite and thirst is that the amount of salt ingested generally overestimates the sodium deficit. This latter difference between water and salt intake is best illustrated by the natural stimulus for sodium appetite: dietary sodium deprivation. In rats, only 1–2 mmol of sodium are lost during the first 1–2 days of sodium deprivation, followed by near-total urinary retention as aldosterone levels increase (Contreras & Hatton, 1975; Stricker et al. 1991; Lane et al. 1997). Nonetheless, sodium deprivation for an additional 6–7 days stimulates as much salt intake as other manipulations that produce much larger volume deficits (see Fig. 8 of Scheidler et al. 1994). Thus, while sodium appetite is an appropriate response to hypovolaemia in a teleological sense, its magnitude is not always proportional to the original decrease in extracellular fluid volume.

What, then, are the physiological signal(s) that instruct the brain to seek and ingest sodium? This important issue remains unsettled, and many proposed answers remain controversial. Figure 4 highlights a number of the stimulatory and inhibitory factors discussed below.

View larger version (27K): [in this window] [in a new window]   Figure 4.  Various stimulatory and inhibitory signals act on the brain to regulate sodium appetite See text for details, discussions and references regarding evidence in support of each mediator.

Although Richter's original experiments with adrenalectomy-induced sodium appetite showed that this hormone is not necessary for sodium appetite, he subsequently discovered that treatment with high-dose adrenal mineralocorticoids stimulated the ingestion of large volumes of saline, even in control animals (Rice & Richter, 1943). In low doses that merely restore baseline renal sodium conservation, the administration of an adrenal mineralocorticosteroid (aldosterone or others, such as deoxycorticosterone) reduces sodium appetite in an adrenalectomized animal (Wolf, 1965; Fregly & Waters, 1966; McEwen et al. 1986; Tordoff et al. 1993). Higher doses, however, robustly stimulate the ingestion of large amounts of saline in both adrenalectomized and adrenal-intact rats (Wolf, 1965; Fregly & Waters, 1966).

The ingestive behaviour stimulated by mineralocorticoids is uniquely specific for sodium. In sharp contrast to other stimuli, such as angiotensin II, mineralocorticoids have little or no effect on water intake (Braun-Menendez, 1950; Wolf et al. 1974; Vallon et al. 2005; Geerling & Loewy, 2006c). Unlike the less-than-straightforward pharmacological manipulations required to increase salt intake using systemic infusions of AII (Fitts & Thunhorst, 1996; reviewed by Weisinger et al. 1996), there is no ambiguity regarding the robust ability of systemic mineralocorticoid administration to selectively increase sodium appetite in rats. Unfortunately, this stimulation requires repeated administration of large and generally supraphysiological doses over several days. Acute administration of physiological doses of aldosterone only produces a small and variable increase in salt ingestion (Wolf, 1964b; Wolf & Handal, 1966).

Therefore, although the specificity of aldosterone for stimulating salt intake is unique, its physiological role probably lies more in gradually boosting sodium appetite, and not as an acute stimulus for this behavioural state. An important role has been demonstrated for the activation of mineralocorticoid receptors (MRs) in the brain (Sakai et al. 1986; Francis et al. 2001; Sullivan et al. 2004), although saline intake after diuresis was not reduced by complete inhibition of adrenal steroid synthesis (Rowland & Morian, 1999). The effects of aldosterone are magnified greatly by a concurrent elevation in adrenal glucocorticoids [especially corticosterone, a combined MR and glucocorticoid receptor (GR) agonist that readily enters the brain; Wolf, 1965; Ma et al. 1993; Zhang et al. 1993] or by an intracerebral injection of the peptide hormone angiotensin II (Fluharty & Epstein, 1983). Also, the unique specificity of mineralocorticoids for salt ingestion led to the identification of a unique group of aldosterone-sensitive neurones in the brainstem, which are activated specifically in association with sodium appetite (Geerling et al. 2006a; discussed below, under the subheading Where is sodium appetite regulated in the brain?).

Angiotensin II.  During hypovolaemia, an elevation in circulating AII is an important stimulus for thirst (Fitzsimons, 1998). Like thirst, circulating AII production increases rapidly in response to hypovolaemia. Following the release of renin from cells in the juxtaglomerular apparatus of the kidney, AII is generated by the enzymatic cleavage of circulating angiotensinogen (produced in the liver) to angiotensin I (AI), which is further processed to AII by angiotensin-converting enzyme (ACE). AII plays an important role in the stimulation of sodium appetite, but does not, on its own, selectively stimulate the ingestion of salt relative to water. Many details regarding the contribution(s) of AII to the stimulation of sodium appetite remain a matter of controversy.

Direct infusion of AII into the brain rapidly stimulates a large increase in water intake, and increases the ingestion of saline (Buggy & Fisher, 1974; Avrith & Fitzsimons, 1980). In sharp contrast to the selective increase in salt intake that is stimulated by dietary sodium deprivation (or high-dose mineralocorticoid administration), however, AII stimulates a high ratio of water-to-saline intake (see, for example, data of Prakash & Norgren, 1991). To reconcile this discrepancy with a possible role for AII in sodium appetite, it has been suggested that AII separately influences thirst versus sodium appetite via the differential activation of separate intracellular signalling cascades under various physiological conditions (Daniels et al. 2005). Also, as discussed in a subsequent subsection (The disinhibition hypothesis), larger increases in AII-stimulated saline intake have been obtained after the inhibition of central oxytocin release (Blackburn et al. 1992a; but see Fitts et al. 2003), or after the injection of various neurotransmitter antagonists into a viscerosensory region in the brainstem that mediates the viscerosensory feedback inhibition of ingestive behaviour (Menani et al. 1996; Menani & Johnson, 1998).

In an apparent contradiction to the hypothesis that peripherally generated AII is a key stimulus for sodium appetite during hypovolaemia, intravenous administration of AII rapidly increases water intake, but a substantial increase in saline intake does not occur until many hours later, and may be secondary to the systemic sodium depletion caused by AII-induced pressure natriuresis (Sakai et al. 1990; Yang & Epstein, 1991; reviewed by Weisinger et al. 1996). Also, boosting AII levels immediately after sodium depletion neither accelerates the onset of sodium appetite nor increases the volume of subsequent saline ingestion (Rowland & Morian, 1999; but see Fitts et al. 1985b). Intravenous administration of AII can, however, increase saline intake acutely if rats are first sodium depleted by diuretic administration on the previous day, then repeatedly administered large doses of captopril (to block endogenous AII production), and then infused with AII on the following day (Fitts & Thunhorst, 1996).

In another contradiction to the hypothesis that circulating AII is a critical stimulus for sodium appetite, the inhibition of AII production by an ACE inhibitor such as captopril somehow increases saline ingestion in rats (Evered & Robinson, 1983; Moe et al. 1984). This captopril-stimulated increase in salt intake can be prevented by the concurrent administration of an ACE inhibitor (or AII receptor antagonist) directly into the brain, or by the peripheral administration of extremely large doses of captopril (e.g. 100 mg kg^–1), both of which reduce saline intake after physiological sodium deficiency as well (Elfont et al. 1984; Moe et al. 1984; Weiss et al. 1986). To reconcile these paradoxical findings with the hypothesis that AII is a critical stimulus for sodium appetite, it has been suggested that captopril, by increasing the circulating concentration of unconverted angiotensin I, allows a preferential increase in its conversion to AII in some brain site, presumably a region that is more accessible to blood-borne AI than to captopril, or that produces enough ACE locally to negate inhibition by captopril unless it is given at extremely high doses or infused directly into the brain (Lehr et al. 1973; Evered & Robinson, 1983; reviewed by Thunhorst, 1996).

It is important to note that the increase in sodium appetite (and thirst) stimulated by AII is much greater when it is administered directly into the brain ventricular system after chronic pretreatment with a mineralocorticosteroid (Fluharty & Epstein, 1983; Zhang et al. 1984; Massi & Epstein, 1990; Shade et al. 2002). The doses of AII used to stimulate salt intake generally produce supraphysiological concentrations of this peptide in the cerebrospinal fluid (CSF) and stimulate non-physiological patterns of neuronal activation relative to circulating AII (McKinley et al. 1995).

These findings led to the ‘synergy hypothesis’, which proposes that sodium appetite is the result of simultaneous elevations in peripheral aldosterone and AII produced by the brain's own renin–angiotensin system (Epstein, 1982). In support of this hypothesis, subthreshold doses of aldosterone (given peripherally) and AII (given centrally) synergize to stimulate more saline intake than either hormone alone (Fluharty & Epstein, 1983). Also, simultaneous pharmacological blockade of systemic MRs and AII receptors inside the brain prevented sodium appetite after furosemide diuresis (Sakai et al. 1986). Based upon various findings in diverse experimental models for sodium appetite, however, it remains unclear whether the actions of AII and aldosterone are sufficient for the stimulation of sodium appetite in response to physiological sodium deficiency (Coghlan et al. 1981; Weisinger et al. 1996, 1997a,b,c; Blair-West et al. 1997; Rowland & Morian, 1999), and additional signalling mechanisms are likely to play an important role.

Baroreceptor input.  Since sodium appetite characteristically arises during a sustained reduction in blood volume and is inhibited by hypervolaemia, the peripheral baroreceptor nerves that detect changes in central venous pressure seem well positioned to provide key information. Consistent with this possibility, the salt ingestion stimulated by prolonged hypovolaemia or mineralocorticoid treatment is greatly reduced if the right atrium is distended with a balloon cannula (mimicking the increase in venous return that occurs when blood volume increases), and then rebounds after the cannula is deflated (Toth et al. 1987). Part of this effect could be mediated by atrial or B-type natriuretic peptides, which inhibit salt intake when administered directly into the brain (Fitts et al. 1985a; Antunes-Rodrigues et al. 1986; Blackburn et al. 1995), but this finding also suggests a role for neural input from central venous baroreceptors in the inhibition of sodium appetite.

Whether venous or arterial baroreceptor signalling is relevant for the stimulation of sodium appetite remains unclear. Arterial blood pressure remains well compensated after dietary sodium deprivation or experimental reductions in ECF volume sufficient for the stimulation of sodium appetite (McCance, 1936; Stokes et al. 1986; Webb et al. 1987; Stricker et al. 1994). In fact, some investigators have even found an increase in arterial pressure during chronic dietary sodium deprivation (Webb et al. 1987). Nonetheless, saline ingestion may be potentiated by reduced arterial pressure (Thunhorst & Johnson, 1994). Conversely, sodium appetite was reduced in rats after transection of the nerves that transmit sensory information from arterial baroreceptors (Thunhorst et al. 1994), although no effect was found after destruction of the brainstem region innervated by these nerves (Schreihofer et al. 1999).

Intracerebral sodium concentration.  The sodium concentration of the CSF is directly related to that of the blood plasma (Doi et al. 1992). Available evidence in multiple species indicates that an increased sodium concentration in this fluid compartment not only stimulates thirst (Andersson, 1978), but also inhibits sodium appetite. For example, infusion of hypertonic saline directly into the brain ventricular system consistently reduced the salt intake of sodium-deficient sheep (Weisinger et al. 1979, 1982), and a similar finding was reported in hypovolaemic rats (Chiaraviglio & Perez Guaita, 1986).

Whether a reduction in intracerebral sodium increases sodium appetite remains unclear. Intraventricular sodium-free infusions of certain osmotically active molecules, such as mannitol or sucrose, increased salt intake in sodium-deprived and non-deprived sheep (Weisinger et al. 1979, 1982). Since one of the effects of these infusions was a reduction in CSF [Na⁺], these experimenters suggested that sodium appetite is increased by the activation of low-sodium sensors somewhere in the brain (separate from the high-sodium sensors that increase thirst and inhibit sodium appetite). However, when equivalent reductions in CSF [Na⁺] were produced by infusions of water or a cell-permeable solute (glucose), salt intake did not increase. This discrepancy was interpreted as evidence that cerebral low-[Na⁺] sensors are located somewhere behind the ventricular epithelial barrier (unlike the high-[Na⁺] sensors located in forebrain circumventricular organs, which lack a blood–brain barrier) and therefore only detect shifts in [Na⁺] across this boundary, within the neuropil (see discussion by Weisinger et al. 1982, 1985; Denton et al. 1996).

Both the significance and the location(s) of these hypothetical low-[Na⁺] sensors remain unknown. These findings could not be reproduced in other laboratory species, including rabbits, rats and mice (Denton et al. 1984; Frankmann et al. 1987; Osborne et al. 1990), and no evidence for a low-[Na⁺] sensing mechanism has been detected at the cellular or molecular level. Subsequent findings in transgenic mice, however, did confirm the existence of sensors for high-[Na⁺] in the brain as a unique concentration-activated sodium channel (discussed below, under the subheading Where is sodium appetite regulated in the brain?; Watanabe et al. 2000).

The disinhibition hypothesis.  The lack of evidence for a critical stimulus that increases sodium appetite (in a physiologically relevant model) led to a new school of thought, which suggested that the appetite for salt is stimulated essentially by the same molecular signal(s) as hypovolaemic thirst (primarily angiotensin II), but that it is usually held in check by a dominant inhibitory signal. To explain the extensive delay between the ingestion of water versus salt after the onset of hypovolaemia, this hypothesis invokes one or more central mechanisms that block sodium appetite (but not thirst) until a sufficient amount of water has been ingested. In this model, sodium appetite is gradually released from inhibition as water ingestion produces osmotic dilution of the extracellular fluid (Stricker & Jalowiec, 1970; Stricker & Verbalis, 1987). Although systemic osmotic dilution is not necessary in various conditions known to stimulate sodium appetite (including dietary sodium deprivation), key inhibitory mechanisms have been identified that are consistent with this model.

Initially, central oxytocinergic projections from the hypothalamic paraventricular nucleus were proposed as the primary mechanism for the inhibition of sodium appetite. This hypothesis is supported by a large body of pharmacological evidence showing that salt ingestion was enhanced after central oxytocin blockade by a variety of methods (Stricker et al. 1987; Stricker & Verbalis, 1987; Blackburn et al. 1992a,b, 1993, 1995; reviewed by Stricker & Verbalis, 1996). As mentioned above, salt intake is also inhibited when elevated plasma sodium (hypernatraemia) is detected by specialized cells within the brain (Chiaraviglio & Perez Guaita, 1986; Watanabe et al. 2000), but this inhibitory mechanism appears to operate independently of central oxytocinergic pathways (Blackburn et al. 1995).

Additional inhibitory control over salt intake is mediated by post-ingestive feedback signals that are transmitted by the vagus nerve, through the brainstem and ultimately to the forebrain. These signals play an important role in limiting the amount of salt that a sodium-hungry animal will ingest (Contreras & Stetson, 1981; Curtis & Stricker, 1997; Menani et al. 1998). Further inhibitory mechanisms have been inferred from the attenuation of sodium appetite after intracerebral injection of a variety of neuromodulators, including adrenomedullin (Samson & Murphy, 1997; Samson et al. 1999), atrial natriuretic peptide (Fitts et al. 1985a; Antunes-Rodrigues et al. 1986; Weisinger et al. 1992), cholecystokinin (Menani & Johnson, 1998), neuromedin B (Massi et al. 1988; Flynn et al. 1999), serotonin (Menani et al. 1996, 1998) and somatostatin (Weisinger et al. 1991). It remains unclear where exactly in the brain most of these inhibitory signals operate, or whether any of them must be reduced as part of the stimulation of sodium appetite by a physiological deficit.

Thus, the brain possesses multiple mechanisms for limiting salt intake, but it remains uncertain whether, under physiological conditions, the unique dynamics of sodium appetite are completely explained by disinhibitory control alone. For example, experiments in knockout mice have confirmed a role for oxytocin as an inhibitory modulator of salt intake (Amico et al. 2001; Puryear et al. 2001; Rigatto et al. 2003), but the results did not support the hypothesis that this peptide is individually responsible for gating salt intake in response to stimuli such as AII (see also Polidori et al. 1994; Fitts et al. 2003). Likewise, destruction of the lateral parabrachial nucleus (a key brainstem relay site for ascending post-ingestive signals, in which injections of various neurotransmitter antagonists have been found to increase salt intake) does not appear to potentiate sodium appetite (Johnson & Thunhorst, 1997), despite robustly enhancing thirst (Ohman & Johnson, 1986; Edwards & Johnson, 1991). In fact, sodium appetite is reduced or eliminated, not enhanced, after lesions involving a neighbouring region in the dorsal pons (Flynn et al. 1991; Scalera et al. 1995). These and other findings suggest that sodium appetite is governed by multiple input signals, both stimulatory and inhibitory, that are integrated by the neuronal networks that regulate salt intake.

	Where is sodium appetite regulated in the brain?

Top Abstract Introduction Why do we need... What is sodium appetite... What causes sodium appetite? Where is sodium appetite... Summary Future directions References

 
When a sodium-deficient animal tastes salt, it will excitedly ingest an unusually large quantity (see Supplemental Video). To explain this pronounced behavioural switch, we have proposed a model with three central components (Geerling & Loewy, 2006a), shown in Fig. 5. First, during chronic sodium deficiency, specific groups of neurones provide tonic, increasing signals for sodium need, which motivate salt-seeking behaviour. Second, once salt is tasted, the gustatory apparatus transmits a phasic signal representing sodium detection. Third, these two signals are integrated (along with various inhibitory signals) in one or more forebrain sites that ultimately drive motivated ingestive behaviour.

View larger version (31K): [in this window] [in a new window]   Figure 5.  A general framework of subcortical pathways providing information to the forebrain for the integration of signals related to sodium need with those for gustatory salt detection (in addition to post-ingestive signals that inhibit salt intake) in order to drive the motivated ingestion of salt Details regarding specific brain sites and pathways are provided in the main text.

While the first question remains a matter of debate, the answer to the second question is straightforward: the sensory–integrative–motor network responsible for sodium appetite is not a self-contained reflex pathway in the brainstem. We know that rostral connections must be involved because disconnecting the brainstem from the forebrain eliminates all behavioural evidence for sodium appetite (Grill et al. 1986; Flynn & Stricker, 2003), despite leaving various other hormonal and gustatory–motor reflexes intact (Flynn & Grill, 1983, 1988; Flynn et al. 1995). A critical role for neurones in the forebrain is also supported by evidence that sodium appetite can be eliminated by electrolytic lesions within a dorsolateral region of the hypothalamus (Wolf & Quartermain, 1967), placed in roughly the location of the medial forebrain bundle (a large collection of axonal interconnections between the brainstem and forebrain).

Surprisingly, however, extensive lesions of sensory and motor cortex do not reduce salt ingestion in response to sodium deficiency (Wolf et al. 1970; Wirsig & Grill, 1982). Subregions of medial prefrontal cortex and probably other cortical areas do appear to exert behaviourally relevant influences over salt ingestion (Chiaraviglio, 1984), but these findings indicate that the cerebral cortex is not necessary for the basic regulation of sodium appetite, and that the forebrain neurones primarily responsible for the integrative control of sodium appetite are located within subcortical nuclei.

A wide range of subcortical brain sites have been tested for potential influences on salt intake (see reviews in Wolf et al. 1974; Denton, 1982; Schulkin, 1991; Johnson & Thunhorst, 1997; Daniels & Fluharty, 2004). Rather than providing an exhaustive catalogue of these brain sites, the following summary offers a tentative framework of brain sites specifically linked to sodium appetite in the context of the model described above (their locations within the rat brain are shown in Fig. 6).

View larger version (38K): [in this window] [in a new window]   Figure 6.  Locations of many of the nuclei throughout the rat brain implicated in the regulation of sodium appetite Drawings adapted from the atlas of Paxinos & Watson (2005). Abbreviations: AV3V, preoptic region surrounding the anterior wall of the third ventricle (including but not limited to the OVLT); BST, bed nucleus of the stria terminalis; BSTvl, ventrolateral subdivision of the BST (specifically including the fusiform subnucleus of the BST); CeA, central nucleus of the amygdala; HSD2 neurones, aldosterone-sensitive neurones in the NTS marked by their expression of the enzyme 11β-hydroxysteroid dehydrogenase type 2; LHA-dl, dorsolateral region of the lateral hypothalamic area (the region of effective lesions identified by Wolf and colleagues; see Wolf, 1964a; Wolf & Quartermain, 1967); LPBN, lateral parabrachial region; NTS, nucleus of the solitary tract; OVLT, organum vasculosum of the lamina terminalis; PB, parabrachial nucleus; PBel-i, inner subdivision of the external lateral PB subnucleus; pre-LC, pre-locus coeruleus; PVH, paraventricular nucleus of the hypothalamus; rNTS, rostral (gustatory) region of the NTS; and SFO, subfornical organ.

Interestingly, however, lesions of the SFO, OVLT and surrounding hypothalamic tissue only cause a partial and variable decrease in the stimulation of salt intake by prolonged sodium deficiency. For example, destroying the thirst-critical ‘AV3V’ (anteroventral third ventricular) region around the preoptic recess of the third ventricle (including the OVLT), which consistently abolishes the homeostatic control of water intake, does not eliminate sodium appetite in rats (Johnson & Buggy, 1978; De Luca et al. 1992) and has no detectable effect on sodium appetite in sheep (Weisinger et al. 1993). In some studies, lesions of the SFO did attenuate saline intake (Thunhorst et al. 1990; Weisinger et al. 1990), whereas other investigators found no effect (Andersson et al. 1975; Ruhf et al. 2001; Wilson et al. 2002). Destruction of the both the SFO and the OVLT produces a more consistent reduction in saline ingestion, but does not consistently eliminate it (Fitts et al. 2004).

Conversely, some of the sites interposed within the AV3V region appear to inhibit salt intake. Evidence for this inhibition initially derived from observations in goats and rats that both spontaneous and mineralocorticoid-stimulated saline intake were increased after lesions involving the OVLT and the adjacent median preoptic nucleus (Andersson et al. 1975; Gardiner et al. 1986; Fitts et al. 1990; Fitts, 1991).

Subsequently, Noda and colleagues uncovered a molecular mechanism for the inhibition of salt intake by specialized sodium detectors in the lamina terminalis (Noda, 2006). Many glial cells in the SFO and other parts of the lamina terminalis express a sodium-selective channel, Na_X, which makes them uniquely sensitive to large increases in the extracellular sodium concentration (Watanabe et al. 2000, 2006; Hiyama et al. 2002, 2004; Grob et al. 2004). In mice, Na_X in the SFO is necessary for the rapid stimulation of thirst by hypertonic saline infusion, as well as the inhibition of salt intake after 24 h water deprivation (Watanabe et al. 2000; Hiyama et al. 2004). Also, relative to wild-type mice, Na_X knockout mice over-ingest saline when sodium appetite is stimulated by diuresis and overnight sodium deprivation (Watanabe et al. 2000).

Despite claims that the SFO is ‘essential and sufficient for the control of salt-intake behaviour’ (Noda, 2006), this channel is probably not directly involved in the stimulation of sodium appetite in response to sodium deficiency. The Na_X channel is activated by large increases in the extracellular sodium concentration (greater than 150 mM, as occurs after prolonged water deprivation or after the ingestion of hypertonic saline), but cannot detect a deficit in body sodium content (or concentration). Accordingly, mice lacking the Na_X channel still exhibit an increase in salt intake in response to prolonged sodium deficiency (Watanabe et al. 2000). Interestingly, although rats show a blunting of the rapid thirst response to central infusion of hypertonic saline after lesion of the SFO (similar to Na_X knockout mice), their depletion-induced saline intake is decreased or unchanged, not increased (Thunhorst et al. 1990; Weisinger et al. 1990; Wilson et al. 2002; Fitts et al. 2004), suggesting the existence of both inhibitory and excitatory sensory elements within the SFO. Thus, Na_X-mediated inhibition of salt intake in the SFO may operate indirectly, by inhibiting excitatory output neurones that increase the ingestion of water and salt. This implication is supported by the twofold increased neuronal activation in the SFO of Na_X knockout mice versus wild-type mice after 24 h water deprivation (Watanabe et al. 2000), as well as the Na_X-mediated stimulation of GABAergic interneurones within the SFO in response to hypertonic saline (Shimizu et al. 2007).

Another circumventricular organ essential for the normal control of salt intake is the area postrema (AP). The AP lies atop the brainstem along the caudal floor of the fourth ventricle. Like the SFO and OVLT, it contains many AII receptor-expressing cells (Mendelsohn et al. 1984; Yamada & Mendelsohn, 1989). Unlike the SFO and the OVLT, however, the AP does not stimulate fluid intake in response to AII (Fitts & Masson, 1989). On the contrary, this structure inhibits ingestive behaviour, particularly salt intake (Contreras & Stetson, 1981; Edwards & Ritter, 1982; Curtis et al. 1996). In rats, destruction of the AP leads to a massive increase in the spontaneous ingestion of saline (Contreras & Stetson, 1981; Watson, 1985; Edwards et al. 1993; Wang & Edwards, 1997; Curtis et al. 1999). Interestingly, despite their elevated salt intake, AP-lesioned rats still exhibit a further increase in saline ingestion in response to sodium depletion (Edwards et al. 1993), indicating that the AP is not necessary for the stimulation of sodium appetite.

Thus, a variety of AII-sensitive circumventricular organs exert stimulatory and inhibitory effects on salt intake, none of which are individually necessary for the stimulation of sodium appetite by prolonged sodium deficiency. Collectively, these findings suggest that an important signal for sodium need is generated by cells in one or more brain sites not targeted in previous lesion studies.

One finding from these prior studies hinted that such neurones may be found immediately ventral to the AP, in the nucleus of the solitary tract (NTS). Specifically, salt intake increased very little (or not at all) in rats with AP lesions if collateral damage beneath this structure extended through any more than a limited region of the underlying subpostremal NTS (Contreras & Stetson, 1981; Wang & Edwards, 1997; Curtis et al. 1999). This finding suggests that the inhibitory effect of neurones in the AP may counterbalance a stimulatory signal arising from neurones in the NTS.

Aldosterone-sensitive neurones. Within this region of the NTS, we identified a group of neurones that generate an output signal specifically associated with sodium need. These neurones were originally identified based upon their unusual sensitivity to aldosterone (Geerling et al. 2006a,c). As shown in Figs 7 and 8, they express the mineralocorticoid receptor (MR) and the enzyme 11β-hydroxysteroid dehydrogenase type 2 (HSD2). This enzyme, which inactivates glucocorticosteroids (primarily cortisol in humans; corticosterone in rodents), is necessary for aldosterone sensitivity. This is because glucocorticoids, which bind to the MR with comparable affinity and circulate at 100- to 1000-fold higher concentrations, out-compete aldosterone in most MR-expressing cells. Only cells that express both MR and HSD2 are sensitized to the relatively low levels of circulating aldosterone (Funder et al. 1988; Naray-Fejes-Toth et al. 1998).

View larger version (120K): [in this window] [in a new window]   Figure 7.  Aldosterone-sensitive neurons in the NTS A–C show immunoreactivity for 11β-hydroxysteroid dehydrogenase type 2 (HSD2, stained in black), which labels the aldosterone-sensitive subpopulation of neurones in the rat nucleus tractus solitarius (NTS) as described by Geerling et al. (2006b). Each panel shows a transverse (coronal) section at one of three characteristic rostrocaudal levels through the caudal NTS: just rostral to the area postrema, where large clusters of HSD2 neurones lie along the walls of the fourth ventricle (4V; A); a level containing the area postrema (AP), where the HSD2 neurones form two bands immediately ventral to the dorsal subpostremal NTS (SubPD; B); and a level just caudal to the AP, containing clusters of HSD2 neurones in the commissural NTS (com; C). Each section was counterstained to reveal surrounding Nissl cytoarchitecture (dark blue), and the rostral distance from calamus scriptorius (cs; C) is provided in the upper right-hand corner of each panel. D shows the rostral cluster of HSD2 neurones lining the 4V, as in A, with cytoplasmic HSD2 (green) in combination with nuclear immunoreactivity for the mineralocorticoid receptor (MR, red). Note that the animal used in D was treated with aldosterone to induce MR nuclear translocation (as described by Geerling et al. 2006a). Other abbreviations: Gr, gracile nucleus; T, solitary tract; and X, dorsal motor nucleus of the vagus nerve. Reproduced with permission from Geerling et al. (2006).

View larger version (74K): [in this window] [in a new window]   Figure 8.  The rostrocaudal distribution of aldosterone-sensitive neurones (labelled by immunoreactivity for HSD2, shown in brown) is shown in the nucleus of the solitary tract (NTS) in a dorsal horizontal section (A) and a parasaggital section through the caudal medulla (B) Each section was counterstained to reveal surrounding Nissl cytoarchitecture (dark blue). The HSD2 neurones form a dense rostral cluster, which abuts the fourth ventricle (4V), as shown in A, and extend caudally in an elongated group just dorsal to the dorsal motor nucleus of the vagus nerve (X), as shown in B. Other abbreviations: XII, hypoglossal motor nucleus.

View larger version (65K): [in this window] [in a new window]   Figure 9.  Aldosterone-sensitive HSD2 neurones in the NTS (green) are activated by chronic dietary sodium deprivation and then inactivated once salt is ingested A shows their increased expression of the neuronal activity marker c-Fos (red nuclear immunoreactivity) after 7 days of a sodium-free diet. B, HSD2 neuronal activation (nuclear c-Fos immunoreactivity) is virtually abolished when a sodium-deprived rat is switched to a high-sodium diet. C, the percentage of HSD2 neurones exhibiting nuclear c-Fos immunoreactivity increases progressively over 2–8 days of dietary sodium deprivation, paralleling a similar increase in sodium appetite (Jalowiec & Stricker, 1973; Stricker et al. 1991). Their activation was virtually eliminated in control rats fed the same sodium-deficient chow, then switched to high-sodium chow for 24 h. D, HSD2 neuronal activation is rapidly diminished by salt ingestion, showing a near-total decrease in c-Fos immunoreactivity within just 2 h in salt-hungry rats (8 days of sodium-free diet) that were given a drinking tube containing 3% NaCl solution. Reproduced with permission; see Geerling et al. (2006a) for experimental details.

Based upon their sodium appetite-specific activation, the HSD2 neurones clearly integrate stimuli associated with prolonged sodium deficiency. A confirmed role for the HSD2 neurones in stimulating sodium appetite would explain the previously paradoxical finding that systemic administration of an HSD2 antagonist (allowing unrestricted MR activation by glucocorticosteroids) increases saline intake, despite the simultaneous increase in sodium retention by the kidneys, which should otherwise inhibit sodium appetite (Cooney & Fitzsimons, 1996). Interestingly, a pronounced increase in salt appetite was also reported in a human with impaired HSD2 function (Ingram et al. 1996).

The aldosterone sensitivity, sodium appetite-associated activation and input/output connections of the HSD2 neurones suggest that they play a role in driving sodium appetite. If so, this group of cells could represent an attractive target for pharmacotherapy, especially given the increased blood–brain barrier permeability within this subregion of the NTS (Gross et al. 1990; Broadwell & Sofroniew, 1993). However, these provocative associations do not establish functional involvement, and a causal role for HSD2 neurones in the stimulation of sodium appetite remains to be tested.

Sodium detection: gustatory pathways.  Taste is necessary for the behavioural expression of sodium appetite. Sodium-deficient rats cannot discriminate saline from other solutions when they are infused directly into the stomach (Smith et al. 1968; Mook, 1969; reviewed by Daniels & Fluharty, 2004), or when they include a drug that blocks gustatory sodium channels (Bernstein & Hennessy, 1987; McCutcheon, 1991; Roitman & Bernstein, 1999).

While the precise molecular mechanisms for sodium detection and transduction within the taste buds remain unclear (Chandrashekar et al. 2006), the ascending axonal pathways that convey gustatory input through the brainstem and into the forebrain have been mapped in many neuroanatomy, electrophysiology and lesion studies (Travers et al. 1987). First, information from taste buds located in the tongue and oropharynx is collected by peripheral gustatory nerves (branches of cranial nerves VII, IX and X). While this information is carried by multiple nerves, the main contributor is the chorda tympani branch of cranial nerve VII, which innervates the anterior two-thirds of the tongue and contains the largest complement of sodium-sensitive fibres (Frankmann et al. 1996; Roitman et al. 1999; Blonde et al. 2006). As indicated in Fig. 10, these nerves directly innervate neurones in the rostral one-third of the lateral NTS (distant from the caudal-medial visceroceptive region of the NTS, containing HSD2 neurones below the AP).

View larger version (35K): [in this window] [in a new window]   Figure 10.  Ascending gustatory pathways in the rodent provide information about dietary sources of salt to subcortical nuclei in the forebrain First, gustatory fibres in peripheral branches of cranial nerves VII, IX and X deliver information from taste buds in the tongue and oropharynx to the rostral NTS (rNTS; Travers et al. 1987). Most of the sodium-specific information that is important for the motivated ingestion of salt is transmitted via the chorda tympani branch of cranial nerve VII, which innervates taste buds in the anterior two-thirds of the tongue (Frankmann et al. 1996; Roitman et al. 1999; Blonde et al. 2006). Next, neurones in the rNTS transmit this information rostrally. In rodents, this ascending pathway involves a relay in the caudal parabrachial nucleus (PB), primarily its waist (wa) and medial (m) subnuclei (Norgren & Leonard, 1971; Norgren & Pfaffmann, 1975; Herbert et al. 1990); in primates, gustatory neurones in the rNTS bypass this relay and project directly to the forebrain (see Beckstead et al. 1980). Gustatory relay neurones in the PB project to the forebrain, densely targeting a handful of subcortical nuclei (Norgren, 1976; Bernard et al. 1993; Alden et al. 1994; Karimnamazi & Travers, 1998; Bester et al. 1999). These include primarily: (1) sites within the central extended amygdala complex, p