The transient absorption of 1,3,5-tri-tert-butyl-pentalene in cyclohexane solution has been measured at room temperature exciting at 340 nm with femtosecond pulses. Two relaxation processes have been determined, the first, very rapid, with time constant 650 fs and the second, occurring on a picosecond timescale, with estimated decay time 13 ps. The former is due to the vibronic relaxation from upper S₃ levels to the S₃ minimum, the latter to the decay from S₃ to S₀. This interpretation is in good agreement with results of ab initio calculations on pentalene and its alkyl derivatives, 1,3,5-tri-ethyl, 1,3,5-tri-sec-propyl and 1,3,5-tri-tert-butyl. MCSCF/CAS calculations, perturbatively corrected within the scheme of quasi-degenerate perturbation theory (QDPT), of vertical S₀ → S_n excitation energies allow us to study the dependence of excitation energies on substitution. Calculated S₃ → S_n and S₄ → S_n absorption spectra of pentalene at the S₀, S₃ and S₄ equilibrium geometries indicate that vibronic levels of S₃ rather than of S₄ are excited by the strong pump pulse with subsequent decay within this state.