Overview of 25 parameters measured during 3-hour behavioral assay.

(A) 3-hour timeline of the presentation shown to treated zebrafish larvae. 25 parameters of behavior are measured during 18 periods of visual and acoustic stimuli. (B) Behavioral profiles of treatments using 12 compounds from the Cayman Chemical FDA-approved Drug Library. Each treatment is compared to DMSO-vehicle controls and the resulting differences are color-coded based on an increase (red) or decrease (green) in a particular behavioral parameter. The values represent differences as compared to DMSO controls in percentage points. Treatments are identified by the Cayman Chemical library’s plate number (M1–M11) and well number (A1–H11).

K-means cluster analysis.

876 compound treatments and DMSO controls were assigned to k = 4 clusters. Cluster 4 contains cyclosporine A and 58 compound treatments with similar behavioral profiles.

Behavioral profiles of CsA-like compounds.

Identified 64 compounds that induce behavioral profiles similar to CsA when administered to 5 dpf zebrafish larvae. Each behavioral profile is composed of 25 parameters measuring activity, reactivity, swimming patterns, and optomotor response.

Hierarchical cluster analysis.

(A) Overview of the behavioral profiles elicited by 876 compound treatments. A high-resolution version of this image is included in the supplementary information (Supplementary Figure 2). (B) Cluster of 53 compounds that induce CsA-like behavioral profiles. Red indicates an increase in a behavioral value relative to DMSO controls, while green indicates a decrease. (C) Pearson pairwise correlations of CsA and the 47 CsA-like compounds identified by both K-means and hierarchical clustering.

Predominant targets and pathways perturbed by CsA-like compounds.

(A) Top 10 targets affected by the 876 compounds in the Cayman Chemical FDA-approved Drug Library. (B) Top 10 targets affected by CsA and the 64 CsA-like compounds found in our clustering analyses. (C) Top 25 Wikipathways, Reactome, KEGG, and PANTHER pathways containing targets affected by CsA-like compounds.

IPA analysis showing the predicted effect of CsA-like compounds on Alzheimer’s disease

Orange lines and nodes indicate predicted activation, blue lines and nodes indicate predicted inhibition, and yellow lines indicate inconsistent findings. Red nodes indicate activation through custom input using the Molecule Activity Predictor (MAP). Purple outlines indicate AD-related nodes.

Potential neuroregulatory pathways affected by CsA-like compounds.

We identified 5 major targets of CsA-like compounds: dopamine receptors, adrenergic receptors, calcium channels and related pathways, steroid hormones, and 5-HT receptors. These targets are involved in various processes of both Ca²⁺ and NO homeostasis, which have important roles in regulating neural function. Green arrows: activated pathways. Red arrows: inhibited pathways. Abbreviations: 5-HTR, Serotonin receptor; α₁, alpha-1 adrenergic receptor; β₂, β₂ adrenergic receptor; AC, adenylyl cyclase; Ca²⁺, calcium; CaM, calmodulin; cAMP, cyclic adenosine monophosphate; D, dopamine; DR, dopamine receptor; E, epinephrine; E2, estradiol; eNOS, endothelial nitric oxide synthase; ER, endoplasmic reticulum; GPER, G protein-coupled estrogen receptor; IP3, inositol trisphosphate; NE, norepinephrine; NFAT, nuclear factor of activated T cells; NO, nitric oxide; PKA, protein kinase A; PLC, phospholipase C; PLN, phospholamban; ROS, reactive oxygen species; S, serotonin; SERCA, sarco/endoplasmic reticulum calcium ATPase; VEGFR, vascular endothelial growth factor receptor.