PA is observed among patients with APA and BHA, with about only 5% of cases being familial [36]. However, as in many genetic diseases, rare familial cases provided critical insight into the genetic bases of PA. It has been suggested that currently used classification of familial hyperaldosteronism (FH) into types, FH1–FH4, should only be applied once/if the causative genetic mutation is known [37].

Glucocorticoid-remediable aldosteronism (GRA) was initially described over 50 years ago [38]. Surprisingly, it was observed that aldosterone production could be suppressed by dexamethasone administration. The molecular mechanism was later elucidated in 1992 by Lifton et al. [3940]. Linkage analysis of patients' DNA revealed a fusion of highly homologous genes encoding the two key enzymes required for cortisol (11-β-hydroxylase, cytochrome P450 family 11 subfamily B member 1 [CYP11B1]) and aldosterone (aldosterone synthase, cytochrome P450 family 11 subfamily B member 2 [CYP11B2]) synthesis (Fig. 1). The recombination brings the coding regions of CYP11B2 under control of the regulatory elements of CYP11B1 that are driven by the signaling aimed at glucocorticoid synthesis (i.e., adrenocorticotropic hormone) instead of RAAS AngII and potassium. This results in aldosterone synthase expression in adrenal zona fasciculata rather than zona glomerulosa and ectopic production of aldosterone and hybrid steroids (due to oxidation by both steroid 17-alpha hydroxylase [CYP17A] and aldosterone synthase) [41].

GRA is extremely rare and testing is recommended only for patients with an early onset of PA and in cases of familial PA occurrence or stroke at a young age (<40 years old) [2842]. The early diagnosis of GRA is important because excessive aldosterone production and associated hypertension can be corrected by low doses of dexamethasone [43]. Glucocorticoid suppression of aldosterone was previously used to both diagnose and treat GRA until introduction of the molecular techniques to detect the presence of the chimeric CYP11B1/CYP11B2 gene into the clinic [4445]. MRAs can be used as a second line of therapy to reduce deleterious non-genomic effects of aldosterone [46].

A different type of FH, not suppressible by glucocorticoids, has been described in several affected families with either APA or BAH [474849]. The causative genetic mutation of this type of FH remained elusive for many years. It was suggested that a locus on chromosome 7p22 was segregating with the disease, but only in some and not all cohorts. Targeted sequencing or NGS of the complete locus did not reveal any mutations [50515253]. It was the NGS analysis of the whole exome from one of the original families that uncovered a germline mutation Arg172Gln in the chloride voltage-gated channel 2 (CLCN2) gene (located on chromosome 3q27 instead) that segregated with this disease [495455]. The authors also reported other CLCN2 variants in unrelated patients. CLCN2 mutations appear to be limited to PA diagnosed at early age and it is absent in patients with essential hypertension [5455].

CLCN2 is a member of the CLC voltage-gated Cl⁻ channels. It is broadly expressed. CLCN2 mutations lead to leukodystrophy and infertility, and Clcn2 loss-of-function in mice leads to germ cell and photoreceptor degeneration [56575859]. Germline CLCN2 mutations that associate with PA are gain of function mutations and promote an export of Cl⁻, depolarization of the plasma membrane, and opening of voltage-gated Ca²⁺ channels, with subsequent activation of CYP11B2 transcription [5455].

In the absence of the genetic foundation, this familial form (termed FH2) was thought to be the most prevalent, responsible for 3% to 6% of all PA cases, which, once the genetic basis was established, is considered to be a gross overestimation [60]. CLCN2 mutations were identified in about 10% of cases with young-onset PA without other known germline mutations and in 2% of patients with BAH, which suggests a significantly lower frequency than previously estimated [5455].

A new inherited form of hyperaldosteronism with hyporeninemia, hypokalemia, and severe hypertension was described in several family members, a father and two young daughters, who developed hypertension by the age of 7 [61]. Adrenal glands in the patients showed notable bilateral enlargement and hypertrophy of a cortical compartment that resembled zona fasciculata juxtaposed to an atrophic zona glomerulosa. Intermixing of cells from different adrenal zones was accompanied by appearance of urine 18-oxocortisol and 18-hydroxycortisol steroids [416263]. Unlike GRA (FH1), dexamethasone administration was not effective in suppressing either glucocorticoid or mineralocorticoid synthesis. Hypertension in these patients was refractory to treatment and correction was achieved by bilateral adrenalectomy [61].

Subsequent exome sequencing identified a heterozygous germline mutation in potassium inwardly rectifying channel subfamily J member 5 (KCNJ5) gene located on chromosome 11q24 in the original patients as well as in sporadic PA [64]. KCNJ5 encodes G-protein-activated inward rectifier K⁺ channel 4 (GIRK4, Kir3.4). Interaction with other Kir family members reconstitutes the functional G-protein-activated potassium channel, which controls membrane polarity in the aldosterone-producing zona glomerulosa cells. The mutation identified in the original work results in a loss of K⁺ selectivity and an indiscriminate influx of Na⁺ ions, membrane depolarization and high intracellular Ca²⁺ concentration. Ca²⁺ increase stimulates aldosterone synthesis through the activation of Ca²⁺ signaling pathways [656667]. The original discovery was followed by several other reports of KCNJ5 familial mutations that mostly cluster in or near protein domain responsible for channel selectivity [686970] although mutations in other locations have been documented [71727374]. The clinical characteristics of the FH3 cases vary widely, from mild to severe, in part due to the underlying mutation [7576]. Testing for FH3 is currently recommended only for patients with very early PA onset by peripheral blood DNA sequencing [28]. Treatment for FH3 varies depending on the severity of the disease, with mineralocorticoid antagonists sufficient for treating milder cases and adrenalectomy to successfully resolve resistant hypertension [7277].

Whole exome sequencing analysis identified a recurrent mutation in the calcium voltage-gated channel subunit alpha 1H (CACNA1H) gene in five unrelated patients in a cohort of 40 pediatric patients diagnosed with PA. That type of PA is referred to as FH4 [78] and both germline and de novo mutations of this gene have been reported [79]. Germline CACNA1H mutations have been documented in several disorders including sporadic amyotrophic lateral sclerosis, epilepsy, autism [808182]; however, for most part FH4 patients showed no symptoms that have been ascribed to these disorders [7983].

CACNA1H encodes for the pore-forming subunit alpha 1H of the T-type calcium channel Ca_v3.2 that is expressed in the zona glomerulosa cells [7883] and is activated by depolarizing changes in the membrane potential [84]. The disease presentation varied in patients carrying different mutation, but most mutations are gain-of-function to some degree (although, see [85]) and augmented aldosterone production [787985]. Several patients harbored mutations in Met1549 located in the repeat domain III of Ca_v3.2 that is a part of a conserved methionine-phenylalanine-valine (MFV) motif that is necessary for channel inactivation [86]. Mutations in Met1549 lead to a decrease in the inactivation of the mutant Ca_v3.2 and a prolonged “on” state, leading to an increase in Ca²⁺ influx that augments the expression of CYP11B2. Treatment of HAC15 cells overexpressing Ca_v3.2 Met1549Val mutant channel with a T-type calcium channel blocker prevented CYP11B2 activation and aldosterone synthesis providing the basis for treatment of patients with FH4 [87]. FH4 follows an autosomal dominant pattern of inheritance, but the penetrance is limited as some Met1549 mutation carriers displayed mild or normotensive phenotype, suggesting that other factors limit the impact of gene defect [78]. Patients with CACNA1H have been also found to have developmental disorders and mental retardation [79].

Mutations have also been reported for a gene encoding a Cav1.3 subunit for another channel, CACNA1D, an L-type (high-voltage activated) Ca²⁺ channel expressed in zona glomerulosa [838889]. Similarly to CACNAH1, sporadic alterations in CACNA1D cause gain-of-function and an increase of Ca²⁺ influx with the consequent overproduction of aldosterone [8390]. In some of these patients seizures and cerebral palsy have been documented [9192]. Somatic CACNA1D was reported to be the most frequently mutated aldosterone-driver gene in African-Americans [93].

Besides familial forms of “channelopathies” described, it is well known that somatic mutations found in aldosterone producing tumors such as KCNJ5, CACNA1, ATPase plasma membrane Ca²⁺ transporting 3 (ATP2B3), and ATP1A13 mutations also cause hyperaldosteronism. Mutations in KCNJ5 are considered to be the most common cause of PA overall [74] and estimated to be present in 60% to 70% cases in APA patients of Eastern Asian origin, but only about half of that in Caucasians [697394959697]. Other mutations (e.g., mutations in catenin beta 1 [CTNNB1] and protein kinase cAMP-activated catalytic subunit alpha [PRKACA] genes) lead to a loss of control over cellular proliferation and rise in the number of aldosterone-producing cells [9899100101]. Mutations in the armadillo repeat containing 5 (ARMC5) gene have been reported to cause hypercortisolism in primary bilateral macronodular adrenal hyperplasia (PBMAH) [102]. However, the connection between ARMC5 mutations and PA remains to be better understood [103]. Moreover, it remains to be determined whether somatic mutations leading to augmented aldosterone synthesis occur in an already altered adrenal cortex or they are responsible for both cellular proliferation and aldosterone production. Finally, while it is clear that BAH can be caused by the same genetic mechanisms as APA, the existence of BAH-specific pathways remains an active area of investigations. Mutations in tandem of p-domains in a weakly inward rectifying K⁺ channel (TWIK)-related acid-sensitive K⁺ channels), twik-related acid-sensitive K⁺ channel (TASK-1 and TASK-3) directly affect K⁺ balance in aldosterone-producing adrenal cells and are known to result in autonomous aldosterone production in mouse models [104105106107]; however, adrenal tumors are not formed in these animals and human disease associated with these genes have not been reported. Furthermore, evaluation of normal adrenal cortex specimens in patients without hyperaldosteronism has found that patients with essential hypertension have aldosterone producing cell clusters (APCC). Expression analysis of these APCCs revealed known aldosterone driver mutations: CACNA1D; ATP1A1, found in 35% of APCC's [108]. It is certainly worth considering if APCCs may add to the pathogenesis of hypertension.

Other area of active investigation in hyperaldosteronism explores the involvement of mammalian target of rapamycin complex 1 (mTORC1) pathway in this disease [109110]. mTORC1 is activated in a subset of patients with PA and treatment with the pathway inhibitor, everolimus, decreased BP, and increased renin concentration in patients with PA; a notable reduction of aldosterone levels was observed in some patients [111]. Similarly, tumor necrosis factor α (TNF-α) pathway is implicated in the pathogenesis of this disorder [112]. It has been long postulated that inflammatory cytokines (TNF-α and interleukins [ILs]) can play an important role in the pathogenesis of cardiovascular disease by creating an environment that promotes hypertension [113]. TNF-α and ILs act at the adrenal level in acute or chronic inflammatory states and could significantly stimulate glucocorticoid production [114115]. IL-1 receptor presence has been reported in a patient with an adrenal adenoma and tumor cells responded with cortisol hypersecretion upon IL-1ß treatment [116]. Similarly, expressions of inflammation-associated genes have been reported in APA patients [94]. It is tempting to hypothesize that mTORC and the inflammatory pathways are intertwined, at least in some tumors (Fig. 2) [117]. Our observations in adrenal cortical cell lines derived from patients' tumors confirm the importance of TNF-α signaling in adrenal neoplasia (unpublished data).