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Figure 2. Innovative metabolic assays for brain organoids
Abbreviations: FLIM: Fluorescence lifetime imaging microscopy; MS: Mass spectrometry; ROS: Reactive oxygen species.

offering a deeper understanding of energy utilization and Several factors complicate metabolic studies in
biosynthetic pathways. 125-127 Optogenetics-based metabolic organoids (Table 3). Oxygen and nutrient diffusion present
control further enhances the ability to modulate and study significant challenges, particularly in larger organoids,
metabolic activity with spatial and temporal precision. 128,129 where insufficient oxygen and nutrient availability can
These advancements hold great potential for investigating lead to hypoxic conditions and core nutrient deprivation,
disease mechanisms and developing novel therapeutic skewing metabolic assessments. 133,134 Cellular heterogeneity
strategies in organoid models. further complicates analysis, as neural progenitors primarily
Metabolomics, both targeted and untargeted, provides rely on glycolysis, whereas postmitotic neurons depend on
135
a comprehensive view of the metabolic landscape oxidative phosphorylation. In addition, hyperglycemic
within organoids. Although metabolomics in brain culture media can induce glucose-related stress, increasing
130
organoids is still an emerging field, it offers valuable data reactive oxygen species production and disrupting neuronal
7,136
on key metabolic pathways such as glycolysis, fatty acid differentiation. Variations in glucose and oxygen
metabolism, and oxidative phosphorylation. Integrating concentrations across studies contribute to inconsistencies
131
transcriptomic data with metabolomic analysis enriches in metabolic outcomes. Furthermore, transcriptomic data
the understanding of the metabolic state within organoid alone do not fully capture metabolic function, particularly
models. One promising development is the application enzyme activity, which is critical for understanding
of imaging mass spectrometry to map lipid distributions metabolic processes. Enzyme activity needs to be studied
within human brain organoids, offering new insights into in conjunction with transcriptomic analyses to provide a
132
137
cellular metabolic processes. However, challenges remain, complete metabolic profile.
such as the technical difficulties of spatial metabolomics Addressing these challenges requires optimizing culture
and the integration of these data with transcriptomic conditions, refining metabolic assessment protocols, and
analyses of metabolic enzymes. fostering interdisciplinary collaboration. By overcoming

Volume 1 Issue 3 (2025) 8 doi: 10.36922/OR025100010