Double-labeling immunofluorescence: A simple and effective tool to study pituitary regulation in teleost fish

Abstract

Neuroendocrinology studies are fundamental to understand how external and internal information are integrated within the brain and, via the pituitary signals, allow the homeostasis of organisms. The teleost neuroendocrine system conserved all the major components and functions present in tetrapods, and the adenohypophysis (ADH) is directly innervated by neurosecretory fibers. This taxonomic group represents an excellent model to study the central regulation of vital processes such as reproduction, water balance, and growth, among others. Besides, further characteristics that distinguish teleost’s pituitary are the regional distribution of the different pituitary cell types, the presence of a hormone named somatolactin in cells of the pars intermedia, the lack of pars tuberalis, and two types of gonadotropes, one that produces follicle-stimulating hormone and the other, luteinizing hormone. Within these teleost characteristics, the lack of a canonical median eminence and a direct innervation with different types of associations between nerve terminals and pituitary cells simplifies neuroendocrine studies and allows the use of simple methodologies to understand the central regulation of pituitary hormones. In this chapter, we will first describe the hypothalamic–pituitary axis in teleost fish, and then, we will introduce the basics of double-labeling immunofluorescence as a simple and reliable method to initiate neuroendocrine studies. In addition, we will describe other experimental approaches to further study the effects of different neuropeptides on hormone secretion and receptor localization, which are necessary to complement these kinds of studies. Finally, we will present some examples of how we applied this protocol in our research lines.