As broadly reported in the literature, the free energy cost to induce a trans-membrane pore in a single lipid bilayer is highly dependent on its lipid composition. For membrane fusion, energetics are not only determined by the lipid composition but also by the hydration level of the bilayers for different inter-membrane distances. 8, These effects can be extrapolated to the more complex fusion pore configuration, combining membrane fusion and a water channel formation. Therefore, it is expected that the free energy to form a fusion pore highly depends on the membranes molecular compositions and their inter-membrane separation. Using a stalk-guided fusion pore and continuum model simulations with the SDK forcefield for opposed bilayers at 6.1nm (center-to-center distance), Kawamoto and Shinoda calculated the free energy cost to nucleate a fusion pore between bilayers, with the following results: ∼100 k B T for DOPC:DOPE (1:1), ∼120 k B T for DMPC:DOPE (1:1), ∼200 k B T for DMPC:DOPC (1:1) and ∼250 k B T for pure DMPC. In good agreement, using Gromacs and the Martini 2 coarse-grained model, we have previously reported ∼160 k B T (∼400 kJ/mol) for pure DPPC bilayers and ∼140 k B T (∼350 kJ/mol) for POPC:POPS:POP2 (87.5:10:2.5). Comparatively, the cost to drive a just nucleated fusion pore into a metastable (slightly expanded) configuration is significantly lower. Again, Kawamoto and Shinoda reported ∼30 k B T to stabilize a nucleated fusion pore for DOPC:DOPE (1:1), ∼20 k B T for DMPC:DOPE (1:1), almost no difference for DMPC:DOPC (1:1) and an energy increase of ∼10 k B T for pure DMPC.