Tuberoinfundibular peptide of 39 residues (TIP39) was identified as the endogenous ligand of parathyroid hormone 2 receptor. We have recently demonstrated that TIP39 expression in adult rat brain is confined to the subparafascicular area of the thalamus with a few cells extending laterally into the posterior intralaminar thalamic nucleus (PIL), and the medial paralemniscal nucleus (MPL) in the lateral pontomesencephalic tegmentum. During postnatal development, TIP39 expression increases until postnatal day 33 (PND-33), then decreases, and almost completely disappears by PND-125. Here, we report the expression of TIP39 during early brain development. TIP39-immunoreactive (TIP39-ir) neurons in the subparafascicular area first appeared at PND-1. In contrast, TIP39-ir neurons were detectable in the MPL at embryonic day 14.5 (ED-14.5), and the intensity of their labeling increased thereafter. We also identified TIP39-ir neurons between ED-16.5 and PND-5 in two additional brain areas, the PIL and the amygdalo-hippocampal transitional zone (AHi). We confirmed the specificity of TIP39 immunolabeling by demonstrating TIP39 mRNA using in situ hybridization histochemistry. In the PIL, TIP39 neurons are located medial to the CGRP group as demonstrated by double immunolabeling. All TIP39-ir neurons in the AHi and most TIP39-ir neurons in the PIL disappear during early postnatal development. The adult pattern of TIP39-ir fibers emerge during postnatal development. However, fibers emanating from PIL can be followed in the supraoptic decussations towards the hypothalamus at ED-18.5. These TIP39-ir fibers disappear by PND-1. The complex pattern of TIP39 expression during early brain development suggests the involvement of TIP39 in transient functions during ontogeny.