Slice cultures of the imprinting-relevant forebrain area medio-rostral neostriatum/hyperstriatum ventrale of the domestic chick: Immunocytochemical characterization of neurons containing Ca²⁺-binding proteins

The forebrain area medio-rostral neostriatum/hyperstriatum ventrale, a presumed analogue to the mammalian prefrontal cortex, displays a variety of synaptic changes during auditory filial imprinting. In order to study the underlying basic mechanisms of this synaptic plasticity we developed slice cultures of the medio-rostral neostriatum/hyperstriatum ventrale from newly hatched chicks. As a prerequisite for these investigations and in order to test the suitability of this system for future studies, we performed a thorough characterization of the in vitro tissue, of its cellular components and some of their biochemical features in comparison with in situ tissue. Since in situ the medio-rostral neostriatum/hyperstriatum ventrale has been previously shown to contain three distinct neuron populations characterized by the activity-regulated Ca²⁺-binding proteins parvalbumin, calbindin D28K and calretinin, we used these proteins as neuronal markers to study the survival and preservation of the morphological features of medio-rostral neostriatum/hyperstriatum ventrale neurons in vitro. In agreement with in vivo studies the three Ca²⁺-binding proteins are confined to neuronal cells and they are not colocalized, i.e. they appear to characterize three different neuron populations. The immunoreactive neurons in medio-rostral neostriatum/hyperstriatum ventrale cultures to a certain extent appear to form synaptic contacts with each other, shown by the double immuncytochemical experiments. One difference between cells in vivo and in vitro is their soma size, which is much larger in vitro than in vivo. This and our previous study on neuronal morphology demonstrates that morphologically and biochemically intact neurons can be maintained in medio-rostral neostriatum/hyperstriatum ventrale slice cultures, which may thus provide a suitable in vitro system for further studies of neuronal and synaptic plasticity in vitro.