In this paper we study the global fluctuations of block Gaussian matrices within the framework of second-order free probability theory. In order to compute the second-order Cauchy transform of these matrices, we introduce a matricial second-order conditional expectation and compute the matricial second-order Cauchy transform of a certain type of non-commutative random variables. As a by-product, using the linearization technique, we obtain the second-order Cauchy transform of non-commutative rational functions evaluated on selfadjoint Gaussian matrices.