## 2 Materials and Methods
### Data Extraction  
The selected publications were then divided into six groups based on subject: epileptic network propagation (Table 1), microglia (Table 2), astrocytes (Table 3), neurovasculature (Table 4), neurites (Table 5), and neurometabolism (Table 6). Two types of tables were then generated: study characteristics (see Supplementary Materials: Table S1 Epileptic Network Propagation, Table S2 microglia, Table S3 Astrocytes, Table S4 Neurovasculature, Table S5 Neurittes, Table S6 Neurometabolism) and data extraction tables. The study characteristics table lists specific features of each paper, such as the title, authors, year of publication, animal model(s) used, study design, and the pharmacological agent used to induce experimental seizures. The data extraction tables contain information on the 2 PI technique, including the type of microscope and laser, the excitation wavelength of the laser, the type of objective used and its numerical aperture, and a description of what was visualized. Figure 1: Flow diagram describing the literature search strategy employed. \begin{table}
\begin{tabular}{c c c} \hline
**Title and Authors** & **Microscope** & **Application** \\ \hline  & Bruker & Two-photon calcium imaging of CGAMPRF-positive \\ Role of inhibitory control in modulating focal seizure spread (Liou & T.Sapphire laser (Coherent) & interneurons was performed. Regions of interest were \\  & 940 nm & registered to target CGAMPRF-expressing \\  & 25\(\times\) water immersion objective (1.05 NA) & parvalbumin interneurons. \\ \hline  & [PERSON] & Two-photon calcium imaging was used to visualize the \\ Acute focal seizures start as local synchronizations of neuronal & T.Sapphire laser (Coherent) & activity of cortical neurons in layers / T III of mouse \\ ensembles ([PERSON] et al. [32]) & 940 nm & primary somatosensory cortex in vivo. \\  & 25\(\times\) water immersion objective (1.05 NA) & primary somatosensory cortex in vivo. \\ \hline  & [PERSON] & The activity of cortical neurons was recorded in vivo by imaging changes in fluorescence (OGR-1 or \\  & & 940 nm \\  & 25\(\times\) water immersion objective (1.05 NA) & AAV1-Syn-GCAMPR calcium indicators). \\ \hline  & BioRad Radiance 2000 MP & Two-photon calcium imaging of mouse hippocampal slices \\ Pyramidal cells accumulate chloride at seizure onset ([PERSON] et al. [34]) & T.Sapphire laser (Spectra) & was performed to visualize epileptiform activity in \\  & 20\(\times\) water immersion objective (0.95 NA) & neuronal populations. \\ \hline Fast spiking interneuron control of seizure propagation in a cortical & Pranite Ultima & Two-photon calcium imaging of rat and transgenic mouse \\  & slice model of focal epilepsy ([PERSON] et al. [35]) & 750 nm, 920 nm & cortico-hippemporal slices was performed. \\ \hline  & Pratie Ultima & Two-photon calcium imaging was used to visualize \\  & & T.Sapphire laser (Coherent) & pyramidal cells expressing CGAMPRF 200-250 μm below \\  & & 980 nm & the cortical surface in transgenic mice. \\  & & 20\(\times\) water immersion objective & \\ \hline  & TrIM Stopel II La Vision Biotech & \\ GABAergic inhibition shapes interectral dynamics in awake epileptic & T.Sapphire laser (Coherent) & CCAMPSG was expressed in CA1 hippocampal cells, and \\  & 920 nm & 2 PI of spontaneous intersectral activity was performed \\ mice ([PERSON] et al. [37]) & 16\(\times\) water immersion objective & in vivo over several days to map cellular recruitment. \\  & (1080 NA) & \\ \hline \end{tabular}
\begin{tabular}{c c} \hline Nanoparticle-based fluoroconjugor for analysis of potassium ion & Olympus FV1000 Confocal & Seizures were monitored in brain slices using the \\  & 633 nm & K\({}^{*}\)-sensitive fluoroconjugor F3. \\ \end{tabular}
\end{table} 
Table 1: Epileptic network propagation—data abstraction table catalogues the type of two-photon microscope used and how two-photon imaging (2 PI) was applied in each study within this group of publications.