The physiological characteristics of central neural populations are being increasingly explored in slice preparations. A major challenge of this approach is to correlate the physiological properties of individual neurones or groups of neurones with their anatomical and chemical properties in order to gain key insights into their functional identities. The present study describes a method for determining the precise topographical position and the immunohistochemical characteristics of neurones in brain slice preparations that are used frequently in electrophysiological investigations. Thick horizontal slices of rat brainstem were re-cut using a method that provided thin sections that were always in the same plane as the parent slice and that were of suitable thickness for immunohistochemistry. Catecholaminergic neurones in these co-planar (horizontal) sections were stained using antisera to tyrosine hydroxylase, the rate-limiting enzyme for catecholamine synthesis. To identify individual catecholamine neurones in the co-planar sections, we constructed a reference atlas of the distribution of catecholamine neurones in the horizontal plane of the rat brain. The combined use of the horizontal atlas and of immunohistochemical techniques in co-planar sections of horizontal slices enables the determination of several key properties: (1) whether a neurone is TH-positive, (2) its precise topographical position and (3) its content of neuropeptides and other immunohistochemical markers. Thus our study offers a readily feasible method for correlative anatomy and immunohistochemistry of physiologically identified catecholaminergic neurones in brain slices.