Integration and segregation are accepted as two fundamental principles of brain organization. The brain manages the transitions between different functional states, more segregated or integrated, through neuromodulatory systems. Recently computational and experimental works suggest a pro-segregation effect of cholinergic neuromodulation. Here, we studied the effects of the cholinergic system on brain functional connectivity using both empirical fMRI data and computational modeling. First, we analyzed the effects of nicotine on functional connectivity and network topology of healthy subjects during resting-state conditions and during an attentional task. Then, to find causal mechanisms about these changes, we employed a whole-brain neural mass model embedded into a human connectome for simulating the effects of nicotine. We found that nicotine incremented functional segregation in both empirical and simulated data, and the effects are context-dependent: observed during task, but not in resting-state. In-task performance correlates with functional segregation, discovering a link between functional network topology and behavior. In the model, the drug was modeled decreasing both the global coupling and local feedback inhibition parameters of the model, consistent with the known cellular effects of acetylcholine. Also, we found that the regional density of the nicotinic acetylcholine α4β2 receptor explains better the effects of nicotine at the whole-brain level. Our results confirm that cholinergic neuromodulation promotes functional segregation in a context-dependent fashion, and suggest that this segregation is suited for simple visual-attentional tasks. In addition, we propose biophysical mechanisms to simulate cholinergic neuromodulation in whole-brain models.