The present study is a part of the previously described SYSTEMA project (5, 6) and was conducted from September 2008 through October 2018. Briefly, patients with unexplained syncope or severe orthostatic intolerance were referred to the tertiary syncope unit at Skåne University Hospital in Malmö from hospitals and outpatient care in southern Sweden.

The definition of unexplained syncope was a transient loss of consciousness without an established diagnosis after the initial evaluation according to the current syncope guidelines (7, 8). During the study period, 2,167 consecutive patients (1,316 women, 60.7%; age, 52.6 ± 21.0 years) were enrolled. We identified 248 cOH and 336 dOH patients (27% of the entire cohort); of these 111 with cOH and 152 with dOH had blood samples collected both in supine position and at 3 min of HUT as per the initial SYSTEMA study protocol (2008–2014) (6). The flowchart of the study is illustrated in Figure 1.

Patients were requested to take their regular medications and fast for 2 h before the test but were allowed to drink water. A self-administered questionnaire was used to collect data about past medical history and the characteristics of syncope-related symptoms.

The examination included basic cardiovascular autonomic testing (Valsalva maneuver and active standing) and a standard HUT according to the Italian protocol (9) i.e., a drug-free HUT phase of 20 min or until syncope occurred. If the drug-free phase was negative, 400 μg sublingual nitroglycerin was administered and the patient was monitored for another 15 min. This part of the protocol was reserved for patients with unexplained syncope, in whom the passive phase was inconclusive and standing systolic BP was over 90 mmHg. The hemodynamic response during the drug-potentiated HUT phase was not a part of OH evaluation. Classical OH was defined as a sustained decrease in SBP ≥20 mmHg or DBP ≥10 mmHg during first 3 min of head-up tilt (HUT), whereas dOH as SBP/DBP fall ≥20/10 mmHg occurring first after 3 min of tilt testing (10), excluding the obvious pattern of vasovagal reflex i.e., typical prodrome and bradycardia preceding or coinciding with a significant BP fall. A pathologic Valsalva maneuver was defined as the absence of an increase in heart rate during phase II, absence of late phase II blood pressure recovery and delayed blood pressure recovery without the characteristic overshoot during phase IV (7).

All disputable or difficult to interpret cases were resolved by adjudication involving one syncope and autonomic expert, and the examining physician.

Beat-to-beat blood pressure and electrocardiogram were monitored continuously by a validated non-invasive method (Nexfin monitor; BMEYE, Amsterdam, Netherlands or Finapres Nova, Finapres Medical Systems, PH Enschede, Netherlands) (11, 12).

Blood samples were collected from an intravenous line, both in supine position and at 3 min of HUT. The decision to collect blood at 3 min of HUT was based on previous studies (13) indicating that the central blood volume displacement to the lower body, heart rate, and total peripheral resistance reach a steady state at 3 min of orthostasis, which implies that the initial neuroendocrine responses are then fully developed.

Measurement of plasma neuroendocrine concentrations has previously been described (6). In brief, we analyzed blood samples in supine position and at 3 min of HUT concentrations of epinephrine, norepinephrine, and non-active peptides C-terminal-pro-arginine-vasopressin (CT-proAVP), C-terminal- endothelin-1 (CT-proET-1), mid-regional-fragment of pro-atrial-natriuretic-peptide (MR-proANP), and mid-regional-fragment of pro-adrenomedullin (MR-proADM). The non-active peptides are generated from the pro-neuroendocrine molecule in ratio 1:1 to the active neuropeptide and are more stable and suitable for analysis compared with their corresponding biologically active substances. High-performance liquid chromatography with fluorescence detection method was used for epinephrine and norepinephrine (14). For measurement of CT-proAVP, CT-proET-1, MR-proANP, and MR-proADM, ThermoScientific BRAHMS assays (BRAHMS GmbH, ThermoFisher Scientific, Neuendorfstrasse 25, 16761 Hennigsdorf, Germany) were used according to the manufacturer's instructions (15, 16).

The regional ethical review board in Lund, Sweden approved the study protocol (reference no 82/2008). All study participants gave written informed consent.

The main characteristics of the study population are presented as mean and standard deviation for normally distributed continuous variables, as median and interquartile range for non-normally distributed variables and percentages for categorical variables. Intergroup differences were analyzed using ANOVA test for non-categorical variables and Pearson's chi-square test for categorical variables. Neuroendocrine concentrations were log-transformed and standardized (expressed per 1 standard deviation). Logistic regression model adjusted for age and gender was applied to test inter-group differences in neuroendocrine concentrations.

Statistical analyses were carried out using IBM SPSS Statistics version 25 (SPSS Inc., Chicago, IL, USA). All tests were two-sided and p < 0.05 was considered significant.

Consistent with previous reports, we found that patients with dOH were younger than those with cOH, supporting the idea that dOH might be an early presentation of autonomic dysfunction, preceding cOH (29). Another finding that supports this is that cOH patients more often had a pathologic Valsalva maneuver than dOH patients. It has been reported that cOH is associated with more severe autonomic dysfunction compared with dOH (30). Conditions that that are associated with autonomic dysfunction and OH, such as Parkinson's disease (31, 32) and reduced renal function (33, 34), were also more common in the cOH group. Surprisingly, diabetes, which is a common cause of secondary OH, was equally prevalent in the two groups.

Orthostatic hypotension is frequently associated with supine hypertension (35, 36). Goldstein found that OH was associated with supine hypertension in patients with primary neurodegenerative diseases. These patients also had low plasma concentrations of norepinephrine, indicating that the supine hypertension in OH is caused by other mechanisms than the hyperactivation of the sympathetic nervous system (35). The observation that cOH patients more frequently have supine hypertension implies that cOH is associated with more severe abnormalities of both autonomic and neuroendocrine control mechanisms.

Studies have shown that cOH is associated with increased risk of cardiovascular disease (3, 37), which can explain the higher incidence of pacemaker treatment in cOH patients. Autonomic dysfunction with sympathetic denervation of the heart that caused sick sinus syndrome has been reported (38). Cardiac denervation could be a possible explanation for the observation that cOH patients reported less palpitations before syncope.

Delayed OH was first described by Streeten and Anderson (4) and has been viewed as a benign condition, but more recent studies indicate otherwise. Gibbons and Freeman followed individuals diagnosed with cOH and dOH for 10 years and discovered that 54% of patients with dOH eventually developed cOH, and 31% developed alpha-synucleopathies (39). Mortality in dOH was increased compared with controls, but not as high as in cOH. Such long-term data indicate dOH not to be a benign non-progressive condition, but in the majority of cases, is an early presentation of cOH.

Better characterization of classical and delayed orthostatic hypotension can have implications for diagnosis, prevention, and treatment of these conditions. In clinical practice, delayed orthostatic hypotension is often overlooked although there are similar clinical consequences in both variants such as syncope as indicated by this study. Whether delayed and classical orthostatic hypotension represent a sequential worsening of the same disease, or constitute two different entities of cardiovascular autonomic dysfunction with different therapeutic approaches, warrants further research.

AF reports personal fees from Medtronic Inc. and Biotronik outside the submitted work; RS reports personal fees and other from Medtronic Inc., St. Jude Medical Inc. (Abbott Laboratories) outside the submitted work; RS is a member of the speaker's Bureau St. Jude Medical/Abbott Inc.; RS is shareholder in Boston Scientific Inc., Edwards Lifesciences Inc. and AstraZeneca PLC. The remaining authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.