Blood-brain barrier (BBB) penetration by aldosterone is substantially less than by most other steroid hormones, as first demonstrated by Pardridge and Mietus (136, 138). Using the Oldendorf technique (132), they observed that reduced brain uptake of certain steroid hormones was associated with their increased molecular polarity, as reflected by the hydrogen bonding number, as shown in A, where n is the sum of 2 per -OH (hydroxyl) group and 1 per =O (ketone) group (in solution, aldosterone exists in equilibrium between 2 states: a predominant hemiacetal with n = 6 and an 18-aldehyde with n = 7). B: only a small fraction of labeled aldosterone (n = 6–7) or cortisol (n = 8) enters the brain, whereas corticosterone (the primary glucocorticoid in rodents; n = 6) exhibits substantial penetration of the BBB. “Brain uptake index” of 100% represents BBB penetration of a freely diffusible reference molecule, butanol (138). C: brain uptake of aldosterone [disintegrations per minute (dpm) per gram] was low in every BBB-protected region tested. L, inferior and superior colliculi; T, thalamus and hypothalamus; C, caudate-putamen; H, hippocampus; F, frontal cortex; O, olfactory bulb (136). [Reproduced with permission from Pardridge (135) and Pardridge and Mietus (136, 138).]

In most mineralocorticoid receptor (MR)-expressing cells (A), the primary ligand is corticosterone or cortisol (both designated “CORT”). These glucocorticoids circulate in the blood plasma at 1,000-fold higher concentrations than aldosterone (ALDO). This systemic imbalance between CORT and ALDO is further exaggerated in the brain, at least in rodents, where only a minute fraction of circulating aldosterone penetrates the BBB (49, 136, 138). Thus, in most MR-expressing cells, basal occupation of this receptor by glucocorticoids prevents the binding of aldosterone, which despite an equivalent binding affinity, may be 10-fold more potent in producing transcriptional activation once it is bound to the MR (9). In contrast, cells that express 11β-hydroxysteroid dehydrogenase type 2 (HSD2; B) can convert a large fraction of intracellular CORT to an 11-oxo metabolite with greatly reduced MR-binding affinity. Aldosterone is not a substrate for HSD2, so this mechanism confers cellular sensitivity to physiological concentrations of this hormone. HSD2 is only a partial filter, leaving a significant amount of unaltered corticosterone able to bind the MR (49, 54), probably because of its greater affinity for the MR (K_m <1 nM) than HSD2 (K_m ∼10 nM in rat).

Prominent expression of HSD2 mRNA in a group of cells within the caudal nucleus of the solitary tract (NTS). A and B: autoradiographic in situ hybridization signal for HSD2 mRNA (antisense probe) with dark- and brightfield optics, respectively. C: lack of hybridization signal (sense probe used as a control). [Reproduced with permission from Roland et al. (150).]

A and B: strong hybridization signal in a glial structure known as the subcommissural organ (SCO) with dark- and brightfield optics, respectively. C: weak expression in the caudal, ventrolateral subdivision of the ventromedial nucleus of the hypothalamus (VMHvl). [Reproduced with permission from Roland et al. (150).]

A–C: immunoreactivity for HSD2 (stained in black), which labels the aldosterone-sensitive subpopulation of neurons in the rat NTS, shown as a transverse (coronal) section at 3 characteristic rostrocaudal levels through the caudal NTS. A: just rostal to the area postrema (AP). B: a level containing the AP. C: where clusters of HSD2 neurons lie just ventral and caudal to the AP, spanning the medial (med) and commissural (com) NTS subnuclei. Sections were counterstained to reveal surrounding cytoarchitecture (blue), and distance rostral to calamus scriptorius (cs, in C) is shown at top right in A–C. D: rostral cluster of HSD2 neurons lining the 4V as in A. Note colocalization of cytoplasmic HSD2 (green) with nuclear immunoreactivity for the MR (red). Animal used in D was treated with aldosterone to maximize MR nuclear translocation (as described in Ref. 54). Gr, gracile nucleus; T, solitary tract; X, dorsal motor nucleus of the vagus nerve. [Reproduced with permission from Geerling and Loewy (62).]

Rostrocaudal distribution of aldosterone-sensitive neurons (labeled by immunoreactivity for HSD2, stained brown) in the NTS in a dorsal horizontal section (A) and in a parasagittal section (B) through the caudal medulla. Each section was counterstained (blue) to reveal surrounding cytoarchitecture. HSD2 neurons form a dense rostral cluster abutting the 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. XII, hypoglossal motor nucleus. [Reproduced with permission from Geerling and Loewy (62).]

Within HSD2 neurons in the NTS, nuclear translocation of the MR is sensitive to and largely selective for physiological concentrations of blood-borne aldosterone. Adrenalectomized rats were treated with vehicle, aldosterone (74.4 ± 10.2 ng/dl), corticosterone (7,900 ± 1,400 ng/dl), or corticosterone + aldosterone. For each group, average percentage of HSD2-immunoreactive neurons (green) that exhibited dense, nuclear-filling MR immunoreactivity (red; nuclear MR represents activated receptors that have moved from the cytoplasm into the nucleus after agonist binding) is shown. Both aldosterone-treated groups exhibited a significant elevation relative to groups treated with vehicle or corticosterone alone, and addition of corticosterone to aldosterone did not produce significantly greater MR activation than aldosterone alone. Corticosterone alone produced a small, but significant, increase relative to vehicle. [Reproduced with permission from Geerling et al. (54).]

Aldosterone-sensitive HSD2 neurons in the NTS (green) are activated by chronic dietary sodium deprivation and then inactivated once salt is ingested. A: dramatic increase in percentage of HSD2 neurons expressing the neuronal activity marker c-Fos (red nuclear immunoreactivity) after 7 days of a sodium-free diet. B: virtual elimination of HSD2 neuronal activation (c-Fos immunoreactivity) when a sodium-deprived rat is switched to a high-sodium diet. C: progressive increase in percentage of HSD2 neurons exhibiting nuclear c-Fos immunoreactivity over 2–8 days of dietary sodium deprivation, paralleling a similar increase in sodium appetite (91, 164). Their activation was consistently eliminated in groups of control rats that were fed sodium-deficient chow but then switched to high-sodium chow for 24 h. D: rapid reversal of HSD2 neuronal activation by salt ingestion, resulting in a near-total loss of c-Fos immunoreactivity just 2 h after a drinking tube containing 3% NaCl solution was provided to salt-hungry rats (after 8 days of sodium-free diet). [Reproduced with permission from Geerling et al. (54).]

Aldosterone-sensitive neurons of the NTS (green, HSD2 immunoreactivity) integrate neural input from a wide variety of sources. A: central nucleus of the amygdala (medial subdivision). Axons and boutons labeled after tracer injection (red) densely entwine HSD2 neuronal somata and proximal dendrites. Transverse section. [Reproduced from Geerling and Loewy (56).] B: axons from the paraventricular nucleus of the hypothalamus (magenta) target many neuronal subpopulations within the NTS, including HSD2 neurons. Horizontal section. C: anterograde tracer injections into the AP (red) produce scattered anterograde labeling in the underlying NTS, including axons that target a minority of the HSD2 neurons. Transverse section. [Reproduced from Sequeira et al. (155).] D: filling the nodose ganglion (cell bodies of vagal afferents to the NTS) with an axonal tracer reveals dense projections covering most of the NTS, including a moderate input to HSD2 neurons caudal to the 4th ventricle. Horizontal section. [Reproduced from Shin et al. (159).] E: neurons in the dorsomedial NTS, which produce the neuropeptide neurotensin (magenta), densely innervate HSD2 neurons and other surrounding populations in their subregion of the NTS. Transverse section. [Reproduced from Sequeira et al. (155).]

HSD2 neurons in the NTS project to a limited number of targets, including a pair of relay nuclei in the rostral, dorsolateral pons [the pre-locus ceruleus (pre-LC) and the innermost subdivision of the external lateral parabrachial nucleus (PBel)] as well as a small subregion of the ventrolateral bed nucleus of the stria terminalis (BSTvl). Other, more minor efferent target sites include the A8/A10 ventral tegmental area in the midbrain and the parasubthalamic nucleus in the caudal lateral hypothalamus (for details, see Ref. 57). ac, Anterior commissure; oc, optic chiasm.

HSD2-immunoreactive neurons in the NTS (green) express the nuclear transcription factor Phox2b (red; arrows indicate colocalization). Other Phox2b-immunoreactive nuclei represent non-aldosterone-sensitive NTS neurons that are intermingled with the HSD2 group. Scale bar, 25 μm. [Reproduced with permission from Geerling et al. (53).]

Outside the NTS, HSD2-immunoreactive cells in the adult rat brain exist in the SCO (A) and in the caudal VMHvl (B). All the columnar ependymal (nonneuronal) cells of the SCO are consistently and intensely immunoreactive for HSD2, whereas the VMHvl contains weaker immunoreactivity in a small subgroup of neurons. Scale bar, 200 μm. [Reproduced with permission from Geerling et al. (55).]