Adrenal Function Tests - Short Note

I. Tests for Glucocorticoid (Cortisol) Function

A. Basal Cortisol Measurement

• Cortisol secretion shows diurnal variation: highest at 8 AM, lowest around midnight.
• Loss of this diurnal variation is one of the earliest signs of adrenal hyperfunction.
• Serum cortisol at 8 AM and midnight can be compared; a flat or elevated midnight level is abnormal.
• A random single measurement is generally inadequate due to pulsatile secretion.

B. Dexamethasone Suppression Test (DST)

1. Overnight (Low-Dose) DST - Screening Test for Cushing Syndrome

• Protocol: 1 mg dexamethasone orally at 11 PM; serum cortisol measured at 8-9 AM next morning.
• Normal response: Cortisol suppressed to < 1.8 µg/dL (50 nmol/L)
• Abnormal (positive): Failure to suppress = suggestive of Cushing syndrome.
• False positives: Oral contraceptive pills (OCP - estrogen increases cortisol-binding globulin; OCPs must be stopped 6 weeks before testing), acute illness, depression, alcoholism.

2. Two-Day Low-Dose DST (Confirmatory)

• 0.5 mg dexamethasone every 6 hours for 2 days.
• 24-hour urine free cortisol (UFC) is measured.
• UFC suppressed to < 50% of baseline rules out Cushing syndrome.

3. High-Dose DST (Differential Diagnosis)

• 2 mg dexamethasone every 6 hours for 2 days (total 8 mg) - or overnight 8 mg version.
• Interpretation:
  • UFC suppressed to < 50% of baseline → Cushing disease (pituitary adenoma - ACTH still responds to high-dose suppression)
  • UFC not suppressed + ACTH < 10 pg/mL → Adrenocortical tumor (ACTH-independent)
  • UFC not suppressed + ACTH > 50 pg/mL → Ectopic ACTH syndrome (ACTH > 300 pg/mL in ectopic tumors - resistant to suppression)

C. 24-Hour Urine Free Cortisol (UFC)

• Provides an integrated assessment of cortisol secretion over 24 hours.
• Not affected by conditions that alter serum cortisol levels (e.g., protein binding changes).
• Multiple collections may be needed since patients with Cushing disease can have cyclic hypercortisolism.
• Uses assay-specific upper limit of normal as cutoff.

D. Late-Night Salivary Cortisol

• Normally cortisol reaches a nadir around midnight; Cushing patients lose this nadir.
• Normal (ELISA): salivary cortisol < 145 ng/dL at bedtime.
• Convenient: can be done at home.
• Accuracy similar to 24-hr UFC.
• False positives possible in hypertension and diabetes.

E. CRH Stimulation Test

• Corticotropin-releasing hormone (CRH) is administered IV; ACTH and cortisol responses are measured.
• Useful in combination with DST to differentiate pituitary Cushing disease from ectopic ACTH syndrome.
• Pituitary adenomas typically show an exaggerated ACTH/cortisol response to CRH; ectopic tumors do not.

F. Inferior Petrosal Sinus Sampling (IPSS)

• Gold standard for localizing ACTH source when MRI is negative or equivocal.
• Central-to-peripheral ACTH gradient > 2 (basal) or > 3 (post-CRH) confirms pituitary source.

II. Tests for Adrenal Hypofunction

A. Short (Rapid) Cosyntropin Stimulation Test

• Rationale: ACTH (cosyntropin = synthetic ACTH) stimulates the adrenal cortex to produce cortisol. Patients with adrenal destruction (Addison disease) show no or inadequate cortisol rise.
• Protocol:
  1. Baseline morning cortisol drawn.
  2. Cosyntropin 250 µg given IM or IV.
  3. Cortisol measured at 30 and 60 minutes.
• Normal response: Peak cortisol ≥ 18-20 µg/dL (500-550 nmol/L), or a rise (Δ) of 7-10 µg/dL. (The peak value is more diagnostic than the delta.)
• Abnormal: Inadequate rise suggests primary adrenal insufficiency.
• Note: Patients with pituitary/hypothalamic dysfunction (secondary/tertiary insufficiency) may show a partial rise, as some adrenal function is preserved initially.
• Low-dose variation: 1 µg cosyntropin (more physiologic); slightly greater sensitivity for mild adrenal insufficiency; comparable overall diagnostic accuracy to the 250 µg test.

B. Prolonged (Multi-day) Cosyntropin Stimulation Test

• Cosyntropin infusion over 2-5 days with plasma/urine cortisol measured.
• Useful when chronic ACTH deficiency has caused adrenal cortical atrophy (secondary insufficiency).
• Primary adrenal insufficiency: no rise even with prolonged stimulation.
• Secondary insufficiency: cortisol rises progressively as atrophied adrenal cortex is "re-trained."

C. Insulin Tolerance Test (ITT) / Insulin-Induced Hypoglycemia

• Insulin is administered to induce hypoglycemia (blood glucose < 40 mg/dL), which is a powerful stress stimulus for ACTH and GH release.
• Normal response: Serum cortisol rises to > 18-20 µg/dL.
• Tests the entire HPA axis (hypothalamus → pituitary → adrenal), unlike cosyntropin which only tests adrenal reserve.
• Contraindicated in coronary artery disease, epilepsy, or severe hypopituitarism.
• Considered the gold standard for diagnosing secondary/tertiary adrenal insufficiency.

III. Metyrapone Test

• Metyrapone blocks steroid 11-hydroxylation, inhibiting cortisol and corticosterone synthesis.
• In a normal person, falling cortisol removes negative feedback → pituitary releases more ACTH → adrenal produces more 11-deoxycortisol (cortisol precursor that accumulates).
• Protocol: 300-500 mg every 4 hours for 6 doses; 24-hr urine 17-hydroxycorticoids (17-OHCS) measured before and after. Alternatively, 2-3 g orally at midnight with 8 AM measurement of plasma 11-deoxycortisol or ACTH.
• Normal response: ≥ 2-fold increase in urinary 17-OHCS; 11-deoxycortisol rises in blood.
• Uses:
  • Assesses pituitary ACTH reserve (secondary adrenal insufficiency).
  • In Cushing syndrome: a normal metyrapone response rules out autonomous cortisol-secreting adrenal adenoma/carcinoma (those suppress ACTH and atrophy the normal adrenal cortex).

IV. Tests for Mineralocorticoid (Aldosterone) Function

Primary Hyperaldosteronism (Conn Syndrome) Screening

• Plasma Aldosterone-to-Renin Ratio (ARR):
  • ARR > 20 with plasma aldosterone > 15 ng/dL is suggestive of primary hyperaldosteronism.
  • An aldosterone:PRA ratio > 50 is highly suggestive.
  • Primary hyperaldosteronism: high aldosterone + low renin (volume expansion suppresses renin).
  • Secondary hyperaldosteronism: high aldosterone + high renin.

Fludrocortisone Suppression / Salt Loading Test (Confirmatory)

• Aldosterone should suppress with sodium loading in normal individuals.
• Failure to suppress aldosterone confirms primary hyperaldosteronism.

V. Tests for Adrenal Medullary Function

• 24-hour urine catecholamines and metanephrines: primary screening for pheochromocytoma.
• Plasma free metanephrines: high sensitivity for pheochromocytoma (preferred screening in high-risk patients).
• Normal: urine metanephrines < 1.3 mg/day; plasma normetanephrine < 0.9 nmol/L, plasma metanephrine < 0.5 nmol/L.

Summary Table