Utility of intracranial EEG networks depends on re-referencing and connectivity choice

Studies of intracranial EEG networks have been used to reveal seizure generators in patients with drug-resistant epilepsy. Intracranial EEG is implanted to capture the epileptic network, the collection of brain tissue that forms a substrate for seizures to start and spread. Interictal intracranial EEG measures brain activity at baseline, and networks computed during this state can reveal aberrant brain tissue without requiring seizure recordings. Intracranial EEG network analyses require choosing a reference and applying statistical measures of functional connectivity. Approaches to these technical choices vary widely across studies, and the impact of these technical choices on downstream analyses is poorly understood. Our objective was to examine the effects of different re-referencing and connectivity approaches on connectivity results and on the ability to lateralize the seizure onset zone in patients with drug-resistant epilepsy. We applied 48 pre-processing pipelines to a cohort of 125 patients with drug-resistant epilepsy recorded with interictal intracranial EEG across two epilepsy centres to generate intracranial EEG functional connectivity networks. Twenty-four functional connectivity measures across time and frequency domains were applied in combination with common average re-referencing or bipolar re-referencing. We applied an unsupervised clustering algorithm to identify groups of pre-processing pipelines. We subjected each pre-processing approach to three quality tests: (i) the introduction of spurious correlations; (ii) robustness to incomplete spatial sampling; and (iii) the ability to lateralize the clinician-defined seizure onset zone. Three groups of similar pre-processing pipelines emerged: common average re-referencing pipelines, bipolar re-referencing pipelines and relative entropy-based connectivity pipelines. Relative entropy and common average re-referencing networks were more robust to incomplete electrode sampling than bipolar re-referencing and other connectivity methods (Friedman test, Dunn-Šidák test P < 0.0001). Bipolar re-referencing reduced spurious correlations at non-adjacent channels better than common average re-referencing (Δ mean from machine ref = -0.36 versus -0.22) and worse in adjacent channels (Δ mean from machine ref = -0.14 versus -0.40). Relative entropy-based network measures lateralized the seizure onset hemisphere better than other measures in patients with temporal lobe epilepsy (Benjamini-Hochberg-corrected P < 0.05, Cohen's d: 0.60-0.76). Finally, we present an interface where users can rapidly evaluate intracranial EEG pre-processing choices to select the optimal pre-processing methods tailored to specific research questions. The choice of pre-processing methods affects downstream network analyses. Choosing a single method among highly correlated approaches can reduce redundancy in processing. Relative entropy outperforms other connectivity methods in multiple quality tests. We present a method and interface for researchers to optimize their pre-processing methods for deriving intracranial EEG brain networks.