Aldosterone is Aberrantly Regulated by Various Stimuli in a High Proportion of Patients with Primary Aldosteronism

Abstract

Context

In primary aldosteronism (PA), aldosterone secretion is relatively independent of the renin–angiotensin system, but can be regulated by several other stimuli.

Objective

To evaluate aldosterone response to several stimuli in a series of patients with PA secondary either to bilateral adrenal hyperplasia (BAH) or unilateral aldosterone-producing adenoma (APA).

Design and setting

Prospective cohort study conducted in a university teaching hospital research center.

Patients

Forty-three patients with confirmed PA and subtyped by adrenal vein sampling (n = 39) were studied, including 11 with BAH, 28 with APA, and 4 with undefined etiology. We also studied 4 other patients with aldosterone and cortisol cosecretion.

Interventions

We systematically explored aberrant regulation of aldosterone using an in vivo protocol that included the following stimulation tests performed over 3 days under dexamethasone suppression: upright posture, mixed meal, adrenocorticotropin (ACTH) 1-24, gonadotropin-releasing hormone (GnRH), vasopressin, and serotonin R4 agonist.

Main outcome measures

Positive response was defined as >50% renin or ACTH-independent increase in plasma aldosterone/cortisol concentration following the various stimulation tests.

Results

Renin-independent aldosterone secretion increased in response to several aberrant stimuli (upright posture, GnRH) in up to 83% of patients with APA or BAH in whom ACTH 1-24 and HT4R agonists also produced aldosterone oversecretion in all patients. The mean significant aberrant responses per patient was similar in BAH (4.6) and in APA (4.0).

Conclusions

Aldosterone secretion in PA is relatively autonomous from the renin–angiotensin system, but is highly regulated by several other stimuli, which contributes to the large variability of aldosterone levels in PA patients.

Primary aldosteronism (PA) is characterized by an excessive secretion of aldosterone despite the resulting sodium retention, volume overload, suppression of renin and angiotensin II, and sometimes hypokalemia (1). The pathophysiology of PA is quite complex and still incompletely understood. During the last decade, novel deoxyribonucleic acid–sequencing approaches combined with immunohistochemistry localization of aldosterone-secreting lesions identified somatic mutations in genes coding for ion channels and ATPases as responsible for up of 90% of aldosterone-producing adenomas (APAs) and germline mutations in the less frequent familial forms of PA (2-6). The pathophysiology of bilateral adrenal hyperplasia (BAH), the most frequent cause of PA, is less well understood; recently, aldosterone-producing cell clusters (APCCs) and BAH specimens were found to carry somatic mutations in CACNA1D (58%) and in KCNJ5 (1%), which suggested that BAH may result from the accumulation of APCCs carrying somatic mutations (7).

Despite these various genetic alterations in PA, aldosterone secretion may not be constitutively autonomous based on studies conducted in a small number of patients demonstrating that aldosterone secretion remained regulated by activation of various aberrant G-protein coupled receptor (GPCRs) in APA and in BAH (8).

It was demonstrated that several aberrantly expressed GPCRs were frequently functional in cortisol-secreting primary bilateral macronodular adrenal hyperplasia or adrenal adenomas, and the possible contribution of these aberrant GPCRs in the pathophysiology of PA was also evaluated. Using a microarray and reverse transcription-polymerase chain reaction in APA samples, overexpression of several GPCRs such as luteinizing hormone human chorionic gonadotropin receptor (LHCGR), 5-hydroxytryptamine (serotonin) type 4 receptor (5-HT4R), GnRH receptor, metabotropic glutamate receptor 3, endothelin receptor type-B like protein (GPR37), and the ACTH receptor (MC2R) were identified (9, 10). The stimulatory effect of 5-HT4R agonists (metoclopramide, cisapride, and tegaserod) on aldosterone secretion was exaggerated in most patients with APA or BAH compared with normal controls (8, 11-13). In accordance, 5-HT4Rs were also overexpressed in most APAs compared with normal adrenals and the normal adjacent adrenal gland (11, 12, 14). Plasma aldosterone levels were reported to increase more after an ACTH bolus in APA than in BAH patients (15, 16) and MC2R was variably overexpressed in most APA tissues studied (12). Other studies confirmed the overexpression of LHCGR in some APA (8) and stimulation of aldosterone secretion following administration of GnRH, hCG, and or human recombinant LH was reported in some patients with PA (12, 17, 18). Aberrant regulation of aldosterone by several GPCR was illustrated in a patient with an APA with aberrant responses to mixed meal, oral glucose, glucose-dependent insulinotropic peptide (GIP) infusion, vasopressin, and tegaserod administration (11). Aberrant GIP receptor (GIPR) expression was confirmed in dispersed cell culture stimulation and in the adenoma by reverse transcription-polymerase chain reaction and immunohistochemistry; overexpression of MC2R and 5-HT4R were also found in this APA (11). Vasopressin type 1 (AVP1R) and 2 receptor (AVP2R) were also shown to be expressed in APA but at variables levels and there was no correlation with the vasopressin-induced aldosterone stimulation in patients with APA, BAH, and in controls (12). In dispersed cells of 3/5 APA, thyrotropin (TSH) stimulated aldosterone secretion (19) and thyrotropin-releasing factor (TRH) increased aldosterone secretion in 1 patient with BAH (12); glucagon also increased aldosterone secretion in single cases of APA and BAH (12).

The objective of the present study was to systematically investigate a larger number of patients with PA to examine the prevalence of aberrant regulation of aldosterone by various stimuli in patients with APA or BAH.

Materials and Methods

Patients selection

The study was conducted between 2006 and 2018 in patients in whom diagnosis of PA was made by a 2-step process using criteria recommended by the Endocrine Society guidelines for the diagnosis and management of PA (20). Firstly, patients with indication for PA screening were sampled for an aldosterone to renin ratio (ARR), which was abnormal if the seated ARR was over 550 pmol/L/ng/L/h using plasma renin activity (PRA) or over 110 pmol/L/ng/L using the direct renin concentration. Secondly, PA was confirmed by a plasma aldosterone concentration >277 pmol/L after 4 hours of the NaCl 0.9% infusion suppression test performed in supine posture (SST) or urinary aldosterone >38 nmol/day following 3 days of high oral salt intake (≥200 nmol/day) (20). Every patient with confirmed PA had an abdominal computed tomography to identify the presence of adrenal nodules or hyperplasia.

Forty-three patients with confirmed PA who gave informed consent to participate in this research protocol were included in the study. Forty of them underwent bilateral simultaneous adrenal vein sampling, as previously described, to confirm the source of aldosterone excess (21). Diagnosis of unilateral APA or lateralized dominant disease was made if the basal lateralization ratio (LR) was ≥2 and post-ACTH LR ≥ 4 or if only the basal LR was ≥2 with a basal contralateral aldosterone/peripheral aldosterone suppression ratio ≤1.5 (15). BAH was diagnosed if the basal LR was <2 with post-ACTH LR <4 (21).

In each patient, the 1 mg overnight dexamethasone suppression test (DST) was performed to evaluate possible cortisol cosecretion. The baseline characteristics of the 43 patients with PA included in the study are described in Table 1. Partial results from 7 patients were reported in previous publications (11, 18). A small subset of 4 additional PA patients with cortisol cosecretion (cortisol above 140 nmol/L after 1 mg DST or 4 mg intravenous [IV] DST) accepted to participate in this study and are described in Table 2. Twenty-seven patients with lateralized PA underwent unilateral adrenalectomy based on the adrenal veins sampling (AVS) lateralization. Pathology report and postsurgical outcome are provided in Table 3.

Table 1.

Baseline characteristics of primary aldosteronism patients

Patient #	Sex/Age	Systolic BP (mmHg)a	Diastolic BP (mmHg)a	HypoK (Y/N)	ARR (N < 550 pmol/L/ng/L×h)	1 mg DST cortisol (nmol/L)	Confirmation test		Adrenal CTb (A)denoma (R or L), (B)ilateral lesions or (N)ormal	AVS results				Surgery (Y/N)α
							Urinary aldosterone (N < 38 nmol/d)	SST PAC at 4h (N < 277 pmol/L)		Basal Lateralizationc	Post-ACTH lateralizationβ	Basal CL (Aopp/Ap)	Aldosterone source
1	M/61	163	89	N	7465	20	105		A (R)	2.26	1.40	3.90	Bilateral	N
2	F/66	150	84	Y	4525	23	56		A (L)	4.70	2.72	28.10	Bilateral	N
3	M/66	152	100	Y	3960	93	85		A (L)	6.96	3.22	2.07	Bilateral	N
4	M/34	145	90	Y	669	21	116		N	1.11	1.38		Bilateral	N
5	M/66	129	74	Y	312 (*DRC)	43	95		B	2.34	1.12	1.65	Bilateral	N
6	M/42	143	85	Y	3570	44	61		A (L)	2.62	3.07	13.30	Bilateral	Y (L)
7	M/46	150	95	Y	4190	11	ND	522	N	2.30	1.16		Bilateral	N
8	F/47	127	75	N	2805	33	79		B	1.50	2.16		Bilateral	N
9	M/51	149	92	Y	2225	47	78		B	6.70	2.93	10.18	Bilateral	N
10	F/50	136	85	Y	1616	19	ND	>277	A (L)	5.24	2.65	80.19	Bilateral	N
11	M/45	ND	ND	Y	1125	15	78		A (L)	1.27	1.62		Bilateral	N
12	M/71	ND	ND	Y	1767	59	128		A (L)	ND	ND	ND	ND	N
13	M/64	ND	ND	Y	2050	104	65	202	B	ND	ND	ND	ND	N
14	F/58	122	74	N	1550	18	61		A (L)	ND	ND	ND	ND	N
15	M/62	137	88	Y	2215	28	62		N	ND	ND	ND	ND	N
16	M/36	135	90	Y	4570	40	181		A (L)	20.37	6.11	3.99	Left	Y (L)
17	M/61	200	130	Y	4615	78	>38		A (L)	15.75	9.13	1.24	Left	Y (L)
18d	M/65	158	90	Y	2510	19	107		B	7.28	5.79	1.94	Left	Y (L)
19d	F/34	144	80	Y	3868	28	150		A (L)	143.02	142.89	1.54	Left	Y (L)
20	F/61	185	90	N	4063	32	43		A (L)	5.21	10.86	4.10	Left	Y (L)
21d	F/51	129	79	Y	2343	34	189		A (L)	1.96	9.73	7.00	Left	Y (L)
22d	M/79	160	60	Y	9025	28	80		A (L)	12.72	14.71	1.27	Left	Y (L)
23	M/62	210	95	Y	3105	15	53		N	15.94	13.91	3.71	Right	Y (R)
24	M/68	145	73	Y	14215	43	150		A (R)	7.50	6.00	5.91	Right	Y (R)
25	F/40	150	100	N	733	18	ND	470	A (R)	15.52	3.15	3.13	Right	Y (R)
26d	M/38	150	90	Y	5420	25	150		A (R)	28.38	27.73	1.86	Right	Y (R)
27	M/73	143	71	Y	5220	56	205		B	3.59	3.18	1.28	Right	Y (R)
28	M/59	153	75	Y	2760	38	116		A (R)	37.15	17.49	3.22	Right	Y (R)
29d	F/39	160	90	Y	2073	31	120		A (R)	15.30	9.08	2.36	Right	Y (R)
30	M/58	134	85	Y	3335	24	93		N	45.63	15.66	7.47	Right	N
31	M/49	147	87	N	3180	23	116		B	24.49	6.91	6.26	Right	Y (R)
32	F/51	145	101	Y	339 (*DRC)	12	66		A (R)	74.89	14.36	1.63	Right	Y (R)
33e	M/56	ND	ND	Y	3980	42	ND	324	B	41.09	24.95	1.55	Right	Y (R)
34	M/66	140	85	Y	4260	19	51		N	41.43	9.70	1.41	Right	N
35	M/55	132	82	Y	128 (*DRC)	76	ND	503	B	7.59	5.28	2.95	Right	Y (R)
36	M/60	144	90	Y	4165	38	112		A (R)	47.85	12.17	2.83	Right	Y (R)
37	M/56	145	90	Y	3055	28	83		B	8.91	5.94	6.59	Left	Y (L)
38	M/67	120	73	Y	1525	27	49		A (L)	39.59	9.67	2.34	Left	Y (L)
39	M/52	120	65	Y	987	24	58		B	9.64	9.32	8.80	Right	Y (R)
40	M/41	138	83	Y	285 (*DRC)	30	106		A (L)	3.66	7.18	5.9	Left	Y (L)
41	M/53	150	80	Y	597	20	214		A (L)	4.13	36.39	11.46	Left	Y (L)
42	M/63	150	97	Y	1898	36	145		A (L)	35.36	38.93	3.75	Left	Y (L)
43	M/51	138	95	Y	3700	17	124	514	A (R)	24.66	118.95	5.06	Right	Y (R)
Total (mean ± SD)	75% M/55 ± 12 y

Abbreviations: BP, blood pressure; CL, contralateral suppression; DRC, direct renin concentration; SD< standard deviation; SST, suppression test performed in supine posture.

βdefined as left (L) or right (R) if LR ≥ 4 or as Bilateral if LR < 4.

αindication for unilateral adrenalectomy as defined in our previous publication (21).

Explanation for the variation between treatment and the AVS results in 3 patients:

#6: Considering the age and the predominant L lateralization, patient underwent surgery despite incomplete contralateral suppression (CL).

#30: In the context of a normal adrenal CT and an incomplete CL, it was decided with the patient to use medical therapy.

#34: Patient refused the surgery as there was no adrenal lesion on the CT scan.

^a BP taken during first clinic visit. Patients under treatment with antihypertensive drugs.

^b Adrenal CT scan findings are reported as: A, unilateral adenoma (s); B, bilateral lesions; or N, normal adrenal morphology.

^c Defined as left (L) or right (R) if LR ≥ 2 or as Bilateral if LR <2.

^d 6 patients already reported in (18).

^e Patient already reported in (11).

Table 2.

Baseline characteristics of patients with aldosterone and cortisol cosecretion

Patient #	Sex (M/F)/Age	Elevated BP (Y/N)	HypoK (Y/N)	1 mg DST (nmol/L)	cortisol after 4 mg IV DST (nmol/L)	ARR (N < 550 pmol/L/ng/L×h)	24h UFC (ULN = 330 nmol/d)	Basal ACTH (pmol/L) N: 2–11	Confirmation test		Adrenal CTa Side L or R	Surgery (Y/N)b
									Urinary aldosterone (N < 38 nmol/d)	SST aldo at 4 h (N < 277 pmol/L)
44	F/56	Y	Y	91	383	1171	1052	1,33	ND	1390	B	Y (L1990; R 2004)
45	F/64	Y	Y	ND	407	5500	276	<0.5	106	ND	B	Y (R)
46	F/41	Y	N	ND	543	841	403	0.8	1265	ND	A (L)	Y (L)
47	M/37	Y	Y	495	712	1640	1103	<0.5	136	ND	B	Y (L)

^a Adrenal CT scanning findings are reported as A unilateral adenoma (s), B bilateral lesions or N: normal adrenals.

None of those patients underwent AVS to identify the origin of their PA.

^b The side of the resection in bilateral disease was chosen based on the side with the dominant nodule.

Table 3.

Pathology report and post-surgical outcome in the PA patients in whom unilateral adrenalectomy was performed

Patient #	Pathology report	Clinical improvement at 12 monthsa	Biochemical improvement at 12 monthsa
6	Adenoma	2	1
15	Adenoma	1	1
16	Adenoma	NA	NA
17	Micronodular hyperplasia	NA	NA
18	Adenoma	1	1
19	Adenoma	2	1
20	Adenoma	2	1
21	Adenoma	2	1
22	Adenoma	NA	NA
23	Adenoma	2	2
24	Adenoma	1	1
25	Adenoma	NA	NA
26	Adenoma	NA	1
27	Adenoma	2	NA
28	Adenoma	NA	NA
30	Adenoma	2	2
31	Adenoma	2	1
32	Adenoma	NA	NA
36	Adenoma	2	1
37	Adenoma	2	3
38	Adenoma	1	2
39	Adenoma	2	1
40	Adenoma	2	1
41	Adenoma	2	1
42	Adenoma	2	1
43	Adenoma	NA	NA

Abbreviation: NA, not available.

^a Criteria for clinical and biochemical improvement based on the PASO criteria Williams et al. (22):

1, complete improvement; 2, partial improvement; 3, absence of improvement.

Aberrant in vivo response screening protocol

The investigation protocol was approved by the institutional ethics committee and every patient provided written informed consent. The investigational protocol was described previously (11); patients were administered 1 mg of dexamethasone orally every 6 hours starting 48 hours prior to the first day and during the 3 days of testing to suppress endogenous ACTH effects on steroidogenesis. Angiotensin receptor antagonists and angiotensin converting enzyme inhibitors and beta-blockers were discontinued at least 48 hours prior to testing and mineralocorticoid receptor antagonists were discontinued 6 weeks prior to testing (11). Plasma levels of aldosterone, renin, cortisol, and ACTH were measured at 30-minute intervals for 1 to 3 hours during tests that modulate the levels of ligands for potential aberrant GPCR. All tests were performed fasting and the patient was in the supine position for at least 60 minutes before the tests. The tests were done over a period of 3 days as demonstrated in Fig. 1. On the first day, a 2-hour upright posture test was done, to screen for possible response to elevation of angiotensin II, vasopressin, or catecholamines; this was followed by a standard mixed meal to screen for the presence of GIP or other intestinal hormone receptors and by a 1-24 ACTH 250-μg IV bolus. On the second day, a 100-μg IV GnRH stimulation test to evaluate the response to GnRH, LH, and follicle-stimulating hormone was followed by a 200-μg IV TRH stimulation test (available from 2008 to 2014) to assess the possible contribution of TRH, TSH, or prolactin. On the third day, stimulation by 6 mg of tegaserod or 10 mg of metoclopramide orally (5-HT4 receptor agonists) was done followed by a stimulation with 10 IU of intramuscular arginine vasopressin. Some patients had additional confirmation tests including the response to 2.5 μg of the V2 vasopressin receptor agonist desmopressin administered subcutaneously, to 300 U of human recombinant LH administered IV, to 75 g of oral glucose, to human GIP infused at a rate of 0.6 μg/kg over 60 minutes, or to the infusion of 20 ng/kg of isoproterenol over 30 minutes. An increase in the aldosterone level ≤24.9% was considered negative, an increase between 25% and 49% was considered a partial response, and an increase ≥50% was considered a positive response. Furthermore, to be considered a possible aberrant response, the renin and ACTH levels must have varied by <50% and must be below the lower limit of the normal. The lower limit of the normal for ACTH was 2 pmol/L, 4 ng/L for renin mass, and 0.6 ng/mL/h for renin activity. A positive significant cortisol cosecretion with the agonist stimulation was defined as a cortisol increase by >50% and by an absolute variation of at least 100 nmol/L. In the present cohort, 38 patients underwent the 7 stimulation tests, 4 had 6 stimulation tests (excluding the TRH test as it was unavailable after 2014), and 1 had 5 stimulation tests (missing 5-HT4R agonist and TRH). The effect of endogenous ACTH on aldosterone secretion in PA was evaluated by calculating the relative suppression of aldosterone level following dexamethasone suppression, which was done with aldosterone level at time 0 of the posture test (following supine position for 60 minutes) of day 1 compared with aldosterone level during aldosterone-to-renin ratio during screening for PA (done in sitting position).

Figure 1.

In vivo screening protocol to detect the presence of aberrant regulation of aldosterone secretion in PA.

Assays

Several assays were used to measure the hormones during the period of the study. During all the study period, ACTH was measured with an immunoradiometric assay (ELSA-ACTH, Cisbio Bioassays, Codolet, France) that was associated with total coefficients of variation (total CVs) <10%. Before 2014, plasma renin activity was used and measured with a radioimmunoassay (GammaCoat Plasma Renin Activity, DiaSorin Inc., Stillwater, MN, USA) that was associated with total CVs <9%. After 2014, renin mass was measured with an immunoradiometric method (RENINE III GENERATION Cisbio Bioassays, Codolet, France) that was associated with total CVs <7.5%. During the study period, plasma aldosterone was measured with 3 different radioimmunoassays (latest: DIAsource ImmunoAssays S.A., Louvain-la-Neuve, Belgique) that were associated with total CVs <10%. Cortisol was measured by competitive chemiluminescent immunoassay (Siemens Healthcare Diagnostics Inc., Tarrytown, NY, USA) that was associated with total CVs <8%. LH and follicle-stimulating hormone were measured by chemiluminescent immunometric assays (Siemens Healthcare Diagnostics Inc.) that were both associated with total CVs <8%. TSH and prolactin were also measured by chemiluminescent immunometric assays (Siemens Healthcare Diagnostics Inc.) that were both associated with total CVs <6%. All CVs were obtained from cumulative statistics extracted from our diagnostic laboratory internal quality control programs that included 3 different concentrations for each hormone.

Analysis

We performed a descriptive analysis of the various aberrant responses observed in the PA patients classified with lateralized or bilateral source based on AVS results. We also compared the relative aldosterone increase during ACTH stimulation test and 5-HT4R agonist between the APA and BAH groups using the Mann–Whitney test and a difference between the 2 groups was considered statistically significant if the P < .05.

Results

Baseline characteristics of the patients

A total of 43 patients were included in the isolated PA cohort. The morning cortisol level following the 1-mg overnight DST was <140 nmol/L in all these patients. Thirty-nine patients underwent an AVS to determine the source of aldosterone excess: 11 patients were diagnosed as BAH (patients #1-11) and 28 patients were lateralized with a dominant APA (patients #16-43). Patients #12-15 did not undergo AVS because they declined surgery or because the pretest likelihood of lateralized disease was too low (older age and bilateral adrenal nodules). The PA cohort included 33 men and 10 women (77% male), the median (interquartile range [IQR]) age was 56 (47-63) years old and the prevalence of hypokalemia was 84% (Table 1). Among PA patients, 27 underwent unilateral adrenalectomy and the postoperative outcome are presented in Table 3. We also studied a subgroup of 4 patients (patient #44-47) with overt cortisol and aldosterone cosecretion (Table 2). The morning cortisol following the 4-mg IV DST was between 383 and 712 nmol/L and the basal ACTH was less than 2 pmol/L in all these patients. This subgroup was composed of 1 male and 3 women that were between 37 and 64 years old. Hypokalemia was present in 75% of those patients. Most of them (75%) had bilateral disease based on computed tomography (CT) scan of the adrenals. In patient #46, the left adrenal containing a 3.7 × 2.7 cm adenoma believed to cosecrete cortisol and aldosterone was resected; Cushing’s syndrome was cured; but PA persisted secondary to BAH which was controlled by spironolactone.

Response of plasma aldosterone concentration to different stimulation tests

Among the 11 patients with BAH, a renin-independent aldosterone increase (≥50% of baseline) was observed in 83% during upright posture, 25% following mixed meal, 100% following 250 µg ACTH IV bolus, 83% following vasopressin, 58% following GnRH, 0% following TRH and 100% following a 5-HT4R agonist (Table 4). An isoproterenol stimulation test was done in only 1 patient, with BAH and a positive response to upright posture test, and aldosterone increased only by 39% during the test. One out of 3 BAH patients with a significant aldosterone increase following the mixed meal had an OGTT 75 g that was also associated with a renin-independent aldosterone increase (105%). A recombinant LH stimulation test was performed in 3 out of 7 BAH patients with an aldosterone increase following GnRH, and only 1 patient showed a renin-independent aldosterone increase (64%) (Table 4).

Table 4.

Aldosterone increase (%) with the various stimulation in patients with bilateral disease (BAH) or no AVS done

Patient	2h upright posture test	Mixed meal	ACTH 1–24	Vasopressin	GnRH	TRH	5–HT4 agonist	# response
1	105	–12	1821	103	76 c	–22	182	5
2	296	80 b	1064	160 f	72	–8	1534	6
3	128	–26	392	157 f	71 c	–6	358	5
4	71	–17	1097	130 f	71 c	–14	568 f	5
5	272	100 b	344	219	31	47	533	5
6	70	53 e	941	361 f	22	ND	483	5
7	41	–21	630	305	184 d	47	696	4
8	70	–22	2237	31	245 c	25	344	4
9	17	–12	235	112	10	–4	285	3
10	807	–34	610	111	–12	–34	1837	4
11	458	15	437	135	80 c	3	77	5
Total response	9/11 (82%)	3/11 (27%)	11/11 (100%)	10/11 (91%)	7/11 (64%)	0/10 (0%)	11/11 (100%)
12	126 a	–58	976	229	90 c	–27	369	5
13	44	20	1030	71 f	54 c	24	411	4
14	377	–5	2500	150	39	–31	1382	4
15	373 f	–51	1089	181	41	78	569	5
Total response	3/4 (75%)	0/4 (0%)	4/4 (100%)	4/4 (100%)	2/4 (50%)	1/4 (25%)	4/4 (100%)

Bold = renin independent aldosterone increase >50%.

Abbreviation: ND, test not done.

^a No renin-independent aldosterone to isoproterenol infusion.

^b Not confirmed by GIP infusion or OGTT.

^c Not confirmed by recombinant LH test or no LH test done.

^d Confirmed by LH test.

^e Confirmed by OGTT or GIP infusion.

^f ACTH-independent cortisol increase >50% with at least 100 nmol/L (every test except for ACTH-1–24)

Number in italics are for patients in whom AVS was not performed.

In 28 APA patients, a renin-independent aldosterone increase (≥50% of baseline) was observed in 71% during upright posture test, 11% following mixed meal, 100% following 250-µg ACTH IV bolus, 68% following vasopressin, 54% following GnRH, 17% following TRH, and 81% following a 5-HT4R agonist (Table 5). A GIP infusion test was performed in 1 of the 3 APA patients showing a positive response following a mixed meal and induced a renin-independent aldosterone increase (144%) that confirmed the GIP-dependent aldosterone regulation in that APA patient. A recombinant LH stimulation test was done in 9 out of 15 APA patients with a response to GnRH stimulation and 5 of them responded with a renin-independent aldosterone increase (range 65-167%) during the test (Table 5).

Table 5.

Aldosterone increase (%) with the various stimulation tests in patient with lateralized disease (APA) on AVS

Patient number	2-h upright posture test	Mixed meal	ACTH 1-24	Vasopressin	GnRH	TRH	5-HT4 agonist	# response
16	20	–16	90	69 e	28	5	52	3
17	77	–15	163	85	78 d	11	124	5
18	181	36b	373	131	118 d	7	511	5
19	19	–15	627	7	LHR positive (+ 68%) c	3	13	2
20	619	–30	1140	100	137 d	–100	930	5
21	590	–39	983	193 e	61 c	–45	225	5
22	83	1	235	9	119 c	26	–2	3
23	86	11	2049	119	85 d	–17	1338	5
24	210	–58	7230	116	–31	ND	337	4
25	66	–4	500	82	2	–24	431	4
26	191	29	1100	125	66 c	84	1058	6
27	1385	9	260	8	2	–29	101	3
28	268	42b	244	103 e	28	53	323	5
29	10	–14	484	65	27	–6	175 e	3
30	191	56 b	746	438	347 c	–9	634	6
31	196	–14	105	136	–8	ND	291	4
32	21	–29	238	174	53 d	4	132	4
33	–1	55 a	489	195	–28	31	557	4
34	197	–4	746	31	125 d	–13	112	4
35	50	–5	295	48	10	ND	ND	1
36	7338	34	65	48	–6	10	392	2
37	423	–40	381	107	17	38	141	4
38	31	–13	165	138 e	110 d	65	42	4
39	433	–32	662	76	41	ND	827	3
40	99	11	114	–37	171 d	ND	893	4
41	763	77 b	345	0	90 d	119	276	6
42	182	–25	630	11	146 d	–4	207	4
43	167	–24	411	109	0	–34	471	3
Total response	20/28 (71%)	3/28 (11%)	28/28 (100%)	19/28 (68%)	15/28 (54%)	4/23 (17%)	22/27 (81%)	Mean: 4.0 response/patient

Bold = renin independent aldosterone increase >50%.

Abbreviation: ND, test not done.

^a Confirmed by OGTT and GIP infusion.

^b Not confirmed by GIP infusion or OGTT.

^c Confirmed by LH test.

^d Not confirmed by recombinant LH test or no LH test done.

^e ACTH independent cortisol increase >50% with at least 100 nmol/L (every test except for ACTH 1–24).

In the 4 patients with PA in which AVS was not performed, a significant renin-independent aldosterone increase was observed in 75% during the posture test, 0% following mixed meal, 100% following 250-µg ACTH bolus, 100% following vasopressin, 50% following GnRH, 25% following TRH, and 100% following 5-HT4R agonist (Table 4).

DDAVP stimulation test was done in 9 patients who had responded to upright posture or vasopressin (1 BAH, 6 APA, and 2 patients without AVS) and none had a positive aldosterone increase. Glucagon stimulation test was done in 6 patients (2 BAH and 4 APA), and significant renin-independent aldosterone increase was observed in 2 BAH patients (51 and 68%) (data not shown).

Aldosterone relative increase between BAH and APA patients during ACTH and 5-HT4R agonist stimulation test ( Fig. 2)

Figure 2.

Relative aldosterone increase (%) following ACTH 250 mcg IV bolus and 5-HT4R agonist stimulation.

Following 250-µg ACTH IV bolus, the median (IQR) relative aldosterone increase in BAH group (n = 11) was 714% (403-1089%) compared with 396% (236-725%) in APA group (n = 28). The minimum and maximum aldosterone relative increase were 235% and 2237% in the BAH group, and 65% and 7230% in the APA group. There was no statistically significant difference between BAH and APA but a statistical tendency was observed (P = .06). The effect of endogenous ACTH on the secretion of aldosterone was evaluated indirectly by the relative % of suppression of aldosterone following dexamethasone suppression during at least 48 hours. The median (IQR) relative suppression of aldosterone in BAH was –72% (–44 to –76) and –47% (+2 to –67) in APA. During the 5-HT4R agonist stimulation test, the median (IQR) relative aldosterone increase was 508% (300-1322%) in the BAH group and 291% (124-557%) in the APA group. The minimum and maximum relative aldosterone increase were 77% and 1837% in BAH and –2% and 1338% in APA. A tendency for a difference in relative aldosterone increase was observed between BAH and APA with 5-HT4R agonist stimulation test (P = .07).

Prevalence of multiple aberrant responses in PA patients

All the BAH patients and the patients that did not undergo AVS presented more than 1 significant aberrant response. In fact, the BAH patients had an average of 4.5 aberrant responses per patient on a possibility of 7 tests (excluding glucagon) (Table 4). For the APA group only, 1 patient had only 1 aberrant response which was to ACTH 1-24, but 5-HT4R agonist stimulation was not done in this patient. The prevalence of multiple aberrant response was also high in the APA group with an average of 4.0 aberrant responses per patient from a possibility of 7 tests (excluding glucagon) (Table 5).

Response of plasma aldosterone concentration and cortisol to agonist in the subgroup of cortisol and PA cosecretion (Table 6)

Table 6.

Cortisol and aldosterone increase (%) with the various stimulation tests in patients with aldosterone and cortisol co-secretion

Patient number	2–h posture test		Mixed meal		ACTH 1–24		Vasopressin		GnRH		TRH		5–HT4 agonist
	Cortisol	Aldo	Cortisol	Aldo	Cortisol	Aldo	Cortisol	Aldo	Cortisol	Aldo	Cortisol	Aldo	Cortisol	Aldo
44	6	23	6	–27	356	140	6	21	–21	–17	65	68	78	44
45	52	545	–10	–29	576	459	227	230	5	92	–12	41	–9	97
46	31	238	15	ND	224	439	85	40	39	42	–8	ND	18	605
47	138 a	163 a	6	–23	184	319	176	127	13	19	2	3	49	161

Bold = renin independent aldosterone increase or ACTH-independent cortisol increase >50%.

Abbreviation: ND, test was not done.

^a Isoproterenol infusion associated with a significant renin-independent aldosterone and ACTH-independent cortisol increase.

The systematic screening of aberrant aldosterone and cortisol response was done in 4 patients with clear aldosterone and cortisol cosecretion. All patients had a significant cortisol and aldosterone increase following ACTH 250 µg IV bolus ranging from 184% to 576% and 140% to 459% respectively. In some cases, aldosterone and cortisol significantly increased together with the same stimulus, but 5-HT4R agonist produced an isolated aldosterone increase in 3/4 patients and an isolated cortisol increase in 1/4 patients. GnRH also produced a significant isolated aldosterone increase in 1 patient. The posture test and vasopressin test produced aldosterone and cortisol increase in 2 patients. 1 patient had an isolated aldosterone increase during the posture test and 1 patient had an isolated cortisol increase following vasopressin.

Aberrant response of cortisol to some agonist in PA patients with normal 1 mg DST

A small number of patients with isolated PA (plasma cortisol post 1-mg DST <50 nmol/L) presented a significant ACTH-independent cortisol increase at the same time than their renin-independent aldosterone increase. In BAH patients, 3 had a significant cortisol increase following 1 of the stimuli; 2 following vasopressin only; and 1 following both vasopressin and 5-HT4R agonist (Table 4). In APA, 5 patients presented a significant ACTH-independent cortisol increase, 4 following vasopressin, and 1 following 5-HT4R agonist (Table 5). In the small subgroup of 4 patients that did not undergo an AVS, 1 patient had a significant cortisol increase and it was during the posture test. Six patients with PA had a 1-mg DST cortisol result between 50 and 140 nmol/L (1 BAH, 3 APA, and 2 in which AVS was not done). We defined them as possible subtle cortisol excess (1 mg DST 50-140 nmol/L). Of those patients, only 2 had an ACTH-independent cortisol increase following vasopressin stimulation test (patient #3 and 13).

Discussion

This study demonstrates that aldosterone secretion is regulated by several stimuli that are distinct from the normal Renin-Angiotensin system (RAS) in a high proportion of patients with PA either from lateralized APA or bilateral source (BAH). In this cohort, almost every patient with PA had at least 2 aberrant responses to 1 of the 7 stimulation tests that were performed systematically. The mean number of aberrant responses per patient was 4.6 in BAH, 4.0 in APA, and 4.5 in patients in whom AVS was not performed. The prevalence of multiple aberrant responses was not different between patients with lateralized or bilateral source of aldosterone excess. This suggests that there are pathophysiological similarities between APA and BAH. Nishimoto et al. described cellular clusters of adrenocortical cells that express CYP11B2 without CYP11B1 expression in the subcapsular portion of the adrenal gland (23). These clusters are known as APCCs. Surprisingly APCCs and nodular hyperplasia are also found frequently in portions of adrenocortical tissues adjacent to the dominant APAs (24) and often are found to carry distinct ion channel mutations than the main APA. So, there is often overlap between so called APA and BAH as in lateralized aldosterone excess, there is often a dominant APA but also adjacent nodular hyperplasia often with more than one ion channel mutation being present. In addition to this complex pathophysiology, we do find that despite various driver gene mutations, aldosterone secretion is not autonomous and can be regulated by various levels aberrant receptor expression and complex loops of locally produced ligands for these receptors.

Only 1 previous study tested systematically several aberrant responses in a group of 12 patients with PA (5 APA, 7 BAH) and 8 controls (12). They found increased renin-independent aldosterone secretion in almost every patient with PA following ACTH and metoclopramide (5-HT4R agonist) stimulus. In our cohort, every patient has a larger aldosterone increase following ACTH bolus than the 2-fold increase reported in normal subjects (12). As reported by other groups (12, 25) we also found that aldosterone level decreased by a median of 72% and 47% following dexamethasone suppression in BAH and APA respectively suggesting a functional role of endogenous ACTH on aldosterone secretion in PA; however, a potential confounding effect of different posture between basal and post dexamethasone suppression cannot be excluded. Previous studies identified increased expression of eutopic MC2R mRNA in most APA compared with normal adrenal cortex or nonfunctioning adenoma (12, 26, 27); however, a great variability was reported as MC2R levels were lower than normal adrenals in some, while 1.4- to 20.6-fold overexpression was present in the majority of APA (12). This variability in MC2R expression probably explains the variable response to ACTH bolus and dexamethasone suppression in our cohort. Adrenocortical lesions harboring somatic KCNJ5 mutations tended to have a lower response to ACTH compared with those harboring ATP1A1 or ATP2B3 mutations (28); however, no systematic study has examined the response to ACTH, the expression of MC2R and the somatic mutations in a group of patients with APA or BAH. As few patients with BAH undergo adrenalectomy, MC2R expression was studied in 1 case only and was increased by 20-fold compared with normal adrenal (12). Surprisingly, we found that patients with BAH tended to have a higher relative aldosterone increase following an ACTH 250-µg IV bolus compared with APA. This finding differs from previous studies indicating a larger increase in APA than in BAH (29, 30); this may result from misclassification between patients with single APA and those with adjacent nodular hyperplasia.

Acute administration of serotonin agonists stimulates aldosterone levels approximately 2-fold in normal subjects but with an exaggerated response in a high proportion of patients with PA (31). It was previously demonstrated that perivascular mast cells in the subcapsular region of the adrenal cortex release serotonin that stimulate the release of aldosterone by adjacent APA (32). The effect of serotonin on aldosterone secretion was shown to be mediated 5-HT4R (31). 5-HT4Rs are expressed in normal adrenal cortex but expression was found to be higher but at variable levels in APA, and the response to various 5-HT4R agonists was higher in patients with APA than in normal controls (9, 11, 12, 14, 31). Consistent with these data, only 5 of our patients (11%) with APA did not exhibit an increase in aldosterone levels following intake of 5-HT4R agonist; in the other patients, we found great variability with relative aldosterone increase between 77% and 1837% in BAH and –2% and 1338% in APA without statistical difference between each etiology. 5-HT4R expression was previously reported only in 1 BAH patient and it was more abundant compared with APA (12). The variable response to serotonin agonists between APA and BAH may also result from various subtypes of 5-HT4R variants that are less effective to induce aldosterone production (33).

In vivo administration of GnRH elicited an aldosterone increase in a significant proportion of BAH (58%) and APA (54%) patients from our cohort. A LH-hCG receptor (R)-driven aldosterone secretion was confirmed in 1/3 of BAH and 5/9 of APA following administration of recombinant LH. LH-hCGR was previously found to be expressed in normal adrenal cortex, but was able to stimulate steroidogenesis in vitro only in fetal adrenals (12, 34). LH-hCGR or GnRH receptor (GNRHR) mRNA were overexpressed in a significant proportion of APA at variable levels (as high as 2400-fold) compared with normal adrenal cortex (10). GnRH and hCG were previously reported to stimulate aldosterone secretion in some APA and BAH, but not in normal controls (12, 17, 18). It was suggested that CTNNB1 (β-catenin) somatic activating mutation can drive the expression of LH-hCGR in PA, but we found no CTNNB1 mutations in patients with aldosterone response to LH/GnRH (18, 35). In 1 study, somatic KCNJ5 mutations was negatively correlated with aberrant LH-hCGR and GnRHR expression in APA (36); however, in a limited number of samples, we found no correlation between in vivo response to GnRH/LH stimulus in vivo and presence of KCNJ5 mutations in resected tissues (18). Thus, complex mechanisms regulating gonadal versus adrenal differentiation may result in LH and GnRH receptors and regulation of steroidogenesis in adrenal hyperplasia and tumors (37). Further studies with larger number of samples will be required to better characterize a possible correlation between aberrant receptors, their ligands and somatic genetic alterations.

A significant proportion of BAH (83%) and APA (71%) equally demonstrated a renin-independent response to upright posture in our cohort; this confirms that posture test is not a good test to distinguish bilateral and lateralized etiologies as suggested previously (38). However, it clearly shows that aldosterone secretion is not autonomous in PA but regulated by physiological stimuli in a renin-independent way in contrast with the renin–angiotensin-regulated mechanism in normal subjects. The significant effect of posture on aldosterone secretion was also demonstrated in previous study that demonstrated a high proportion of false negative saline infusion test when the test was done in recumbent position (54.5%) which was less frequent in seated position (10.4%) (39). The renin-independent aldosterone increase in response to posture was suggested to be mostly driven by endogenous vasopressin or epinephrine (β2 agonist receptor) increase as angiotensin II is suppressed (8). One patient with BAH had an isoproterenol stimulation test to evaluate the possible contribution of β2-receptor activation in response to posture test but aldosterone increased only by 39%. One patient with bilateral macronodular hyperplasia (BMAH) had a significant increase of both aldosterone and cortisol following isoproterenol infusion. We observed that in 75% of our PA cohort, a renin-independent aldosterone increase occurred during the upright posture test when beta-blocker or AT1R antagonists had been withdrawn for 48 hours; it is possible that aldosterone response might have been more significant if those drugs were discontinued longer before the test. In future prospective studies it should be interesting to further evaluate the optimal period of withdrawal of beta-blockers or renin–angiotensin system inhibitors to affect response of aldosterone to upright posture. The prevalence of response to vasopressin, probably mediated by vasopressin type 1 receptor (AVPR1, as no response to desmopressin), is similar in our cohort to the response to posture test; 83% in BAH and 68% in APA. However, not every patient with a significant renin-independent aldosterone increase in response to upright posture responded to vasopressin. It is also unclear if the variation of vasopressin induced by upright posture is sufficient to induce a significant aldosterone response as we observed in our cohort (40). In fact, plasma vasopressin concentration in humans are in the pmol/L range and it was reported that a vasopressin concentration in nmol/L are necessary to induce a significant increase in aldosterone secretion in adrenal cells in vitro (41). AVPR1 and AVPR2 are expressed both in normal adrenal and in APA and at similar level of expression (12, 42). However, there was no clear correlation between the levels of expression of AVP1R/AVP2R and vasopressin-induced aldosterone stimulation (12). Vasopressin can induce pituitary ACTH release (43), but this was prevented by dexamethasone administration and confirmed by the absence of ACTH fluctuation during the vasopressin stimulation test in our cohort. No implication of AVP3R has been identified on regulation of aldosterone secretion in primary aldosteronism to date. In our cohort, 9 patients with PA with a response to vasopressin or upright posture, desmopressin stimulation test (agonist for the AVP2R but also for the AVP3R) (44) did not evoke an aldosterone increase. This suggests that AVP3R is probably not the main pathway by which vasopressin induced an increase in aldosterone levels in PA, but further studies with specific AVP1R and AVP3R antagonists will be of interest. It is plausible that vasopressin frequently contributes to aldosterone increases in response to upright posture test in PA, but this hypothesis needs to be validated in future studies using specific antagonists. The possible contribution of catecholamine to aldosterone increase associated with posture in PA has not been thoroughly evaluated by systematic isoproterenol infusion stimulation test or beta blockers in patients with response to posture. In 1 study, administration of an AVPR1 antagonist potentiated the aldosterone response to upright posture in APA, but not in BAH, and it was suggested that this paradoxical effect might be due to sensitization to catecholamines stimulation in some APA (45).

Mixed meal induced renin-independent aldosterone increase in 6/44 (14%) (3 BAH and 3 APA). In one of the 3 APA, which was previously reported, a GIP infusion provoked a significant aldosterone increase (144%) confirming the GIP-dependent aldosterone secretion (11). Increased GIPR expression and aldosterone increase following incubation of dispersed cells with GIP were demonstrated in the aldosteronoma of this patient (11). In our cohort, one BAH had a significant aldosterone increase (105%) after OGTT 75 g but could not be confirmed by GIP infusion. In the 15 PA patients studied by Zwermann et al. none elicited an aldosterone increase in response to mixed meal (12). GIPR was found in normal zona glomerulosa (ZG) cells by immunohistochemistry and a small increase of aldosterone was found following GIP infusion in 3 control subjects (11). Thus eutopic ZG GIPR can be overexpressed in rare cases of PA, while it is not expressed in normal zona fasciculata (ZF) but can be expressed ectopically in some cortisol secreting adenoma (gene rearrangement) or in some cases of cortisol-secreting adenoma with GIP-dependent Cushing’s syndrome (8, 46).

In our cohort, 5/37 (14%) (4 APA and 1 patient without AVS) demonstrated a significant renin-independent aldosterone increase in response to TRH. TRH was already reported to stimulate aldosterone secretion in one patient with BAH and TSH receptor mRNA was reported to be overexpressed in 1 APA (12); none of the normal controls increased aldosterone following TRH administration (12). Another series reported that TSH receptor was overexpressed in APA compared with normal adrenal tissue (19). In dispersed cells of 5 APA, TSH stimulated aldosterone secretion, but also in normal adrenals and there was no correlation between receptor expression and aldosterone response to TSH (19). TRH stimulated both TSH and prolactin release at the pituitary level. It was shown previously in primary adrenal cells culture that prolactin can increase aldosterone concentration in the cell supernatant (47). However, there are no in vivo data that demonstrated the effect of prolactin on aldosterone secretion in APA or BAH. Glucagon was found to stimulate aldosterone secretion by 2 BAH in our cohort. It was previously reported to stimulate aldosterone release by 1 APA, 1 BAH, and 1 normal control (12). However, glucagon receptor was not detected in 15 adrenal glands removed from patients with PA and in 6 normal adrenal (12). The mechanism by which glucagon can stimulate aldosterone secretion in some patients with PA and normal adrenal remains unknown.

We found that some patients with PA and a normal 1-mg DST (<50 nmol/L) or with possible subtle cortisol excess (1-mg DST 50-140 nmol/L) have a significant ACTH-independent cortisol increase concurrently with their renin-independent aldosterone increase. The ACTH-independent cortisol increases were observed mostly following vasopressin stimulation and 5-HT4R agonist stimulation. Vasopressin is known to induce modestly cortisol secretion by the stimulation of AVPR1 in normal adrenal cortex in cell dispersion (not in vivo) and in a high proportion of patients with primary adrenal Cushing’s syndrome, more frequently in primary BMAH (8, 48). The 50% increase in cortisol threshold that we used in this study required a nonequivocal ACTH-independent cortisol increase. So, it seems that the stimuli can significantly increase cortisol secretion in some patients with PA and normal basal cortisol secretion by stimulating the same GPCR that also increases its aldosterone secretion; it probably occurs in ZF-like PA coexpressing CYP11B2 and CYP11B1 with a specific pattern of ion channel mutation (49).

We also evaluated a subgroup of 4 patients with frank cortisol and aldosterone cosecretion and their aldosterone and cortisol responded concomitantly in most cases with the same stimuli. This also suggests that adrenal primary bilateral macronodular adrenal hyperplasia (PBMAH) or adenoma express the ZF and ZG phenotypes and that the aberrant GPCR are functional in CYP11B2 and CYP11B1 expressing cells resulting in aldosterone and cortisol secretion at the same time. However, in some patients there is a response of only aldosterone or cortisol to a stimulus. It is possible that aberrant receptor might be differentially expressed by CYP11B1 and CYP11B2 positive cells; this hypothesis should be explored in future immunohistochemical studies for CYP11B1, CYP11B2 and G-protein coupled receptors on resected adrenal of patients with aberrant response of cortisol or aldosterone to various in vivo stimuli.

Aldosterone secretion is mainly regulated by angiotensin-II and potassium, but can also be stimulated by ACTH and 5-HT4R agonists acting through a different pathway via their G-protein coupled transmembrane receptor activation of the cAMP/PKA pathway leading to an activation of calcium channel and calcium influx to the ZG cells (50, 51). It is likely that the aberrant GPCR which are also normally coupled to the cAMP/PKA pathway also exert their regulation by this signaling system, but this remains to be confirmed by future in vitro pharmacological studies. The molecular mechanisms underlying the response of aldosterone to those ligands and the expression of aberrant G-protein coupled receptor in primary aldosteronism are unknown. In adrenal Cushing’s syndrome some recent mechanisms have been elucidated: rearrangement of the GIP receptor gene was found in a fraction of patients with GIP-responsive adrenal Cushing’s syndrome (46). It is known that adrenal cortex originates from a common progenitor cell population with gonads in the adrenogenital ridge and GATA4 with its cofactors ZFMP2 and SF1 are involved in gonadal development. It is also known that GATA4 is not normally expressed in normal adult adrenal cells. In a case of transient BAH associated with severe Cushing’s syndrome during pregnancy LHCG receptors were shown to be overexpressed and colocalized in undifferentiated subcapsular adrenal cells that was associated with GATA4 upregulation and in ZG cells that was associated with GATA and ZFMP2 upregulation (52). Then, it is possible that some adrenocortical cells can maintain a certain degree of LHCG receptor after their migration from the adrenogenital ridge and thus causing adrenal hyperplasia when stimulated by LH or hCG. Local overexpression of ACTH in bilateral macronodular hyperplasia (53) or of mastocytes and serotonin adjacent to APA often carrying overexpression of 5HT4R (32) were also suggested to be related to maintenance of gonadal progenitor phenotype.

The main strength of this study results from the largest cohort of PA in which aberrant response to 7 tests were systematically studied in vivo under inhibition of ACTH by dexamethasone. Limitations include the lack of a control group and the use of normal response from previous studies (12, 17). The criteria of >50% increase of aldosterone is relatively arbitrary but was confirmed as being clinically significant by other studies and cases reports in PA and adrenal Cushing’s syndrome (8). The main focus of this study being the evaluation of in vivo aberrant responses, molecular confirmation of aberrant GPCRs were not conducted systematically, but they have been reported in part previously (11-18).

In conclusion, aldosterone secretion in PA is independent from the RAS, but regulated by other mechanisms which probably result from complex interactions between diverse ligands and variable aberrant expression of diverse GPCR either in APA or in BAH tissues. This may explain the frequent variations in levels of aldosterone depending on the fluctuation levels of the ligands. We thus propose that the terminology of autonomous aldosterone secretion in PA should be replaced by the terminology of dysregulated aldosterone secretion given that aldosterone secretion in PA varies importantly secondary to fluctuations and interactions between multiple aberrant ligands and their receptors and are not constitutively stable despite the presence of the various ion channel driver mutations.

Glossary

Abbreviations

5-HT4R

5-hydroxytryptamine (serotonin) type 4 receptor

ACTH

adrenocorticotropin

APA

aldosterone-producing adenoma

APCC

aldosterone-producing cell cluster

ARR

aldosterone to renin ratio

BAH

bilateral adrenal hyperplasia

BMAH

bilateral macronodular hyperplasia

CT

computed tomography

CV

coefficient of variation

DST

dexamethasone suppression test

GIP

glucose-dependent insulinotropic peptide

GnRH

gonadotropin-releasing hormone

GPCR

G-protein coupled receptor

LR

lateralization ratio

PA

primary aldosteronism

PRA

plasma renin activity

TRH

thyrotropin-releasing factor

TSH

thyrotropin

ZG

zona glomerulosa

Additional Information

Disclosure Summary: There are no conflicts of interest to disclose.

Data Availability

Some or all datasets generated during and/or analyzed during the current study are not publicly available but are available from the corresponding author on reasonable request.