The hypothalamo-pituitary-adrenocortical axis plays a vital role in adaptation of the organism to homeostatic challenge (J.P.Herman et al., 2003). During most of infancy, from approximately postnatal day 4 to 14, the rat displays a stress hyporesponsive period in the form of markedly attenuated adrenocorticotropin and corticosterone responses to environmental stressors that elicit pronounced elevation of ACTH and corticosterone in pre- and post-stress-hyporesponsive period rats (J.Lehman et al., 2002). After that the pituitary gland undergoes the prominent age-related adaptation changes. The different cell types in the anterior pituitary behave as dynamic populations, as the hypophysis maintains a continuous renewal of cells to ensure a balance between cell division, differentiation, growth arrest and apoptosis (L.Claudius et al., 2006). Numerous discrepancies in the evaluation of activation, hyporesponsiveness, facilitation and dissociation of the hypothalamo-pituitary-adrenocortical axis in the early age may be explained by a very limited information available in the literature on the age-related structural changes in the central link of the axis - adenohypophysis - during early postnatal development (C.Kaur et al., 2002; A.Armario et al., 2004; X.Belda et al., 2004; C.Marquez et al., 2005).
The objective of this study was to evaluate the developmental changes in the pituitary gland of the growing rats during early postnatal development using the modern methods of the quantitative immunohistochemistry.
Preweaning, weaning and early postweaning Sprague-Dawley rats aged 14, 21 and 30 days after birth (1st, 2nd and 3rd age groups accordingly) were weighed and sacrificed by cervical dislocation, their pituitary glands were removed, weighed, fixed in formalin and embedded in paraffin. Serial sections 4 mcm thick were stained with hematoxylin-eosin and immunohistochemically with monoclonal antibodies against ACTH and PCNA using streptavidin-biotin-peroxidase method with subsequent DAB-staining and image-analysis of the immunohistochemically stained slides. Image Pro Plus 4.5 software was utilized to evaluate the volume and numeric density of the immunoreactive cells.
The results obtained in the present investigation demonstrated that the pituitary gland mass increased with age while the relative pituitary gland mass decreased from preweaning to postweaning age, with a difference reaching the level of significance between the 1st and the 3rd age groups (p<0,05).
Routine histological staining demonstrated that in the pituitary gland of the growing rats adenohypopysis and neurohypophysis were clearly demarcated with a distinct subdivision of the adenohypophysis into pars distalis and pars intermedia. The pars distalis was dominated by the chromophobic cells which were distributed in clusters with their nuclei located very close to each other in the groups. Among the chromatophylic cells the share of the oxyphilic cells was higher compared to the basophilic adenocytes, the latter being increased by peripubertal age. Most of the oxyphilic cells were concentrated in the lateral wings of the adenohypophysis while the central part of pars distalis contained comparatively more basophils.
The immunohistochemical staining demonstrated that on the contrary to the hematoxylin-eosin, staining for ACTH revealed concentration of the immunopositive cells in the lateral wings of the hypophyseal anterior part and not in the basophilic center of the pars distalis, though it is known that cortocotrophs are basophilic when stained by hematoxylin eosin. This allowed us to presume that the basophilic staining of the central part of the pars distalis in rats was due to the presence of the other types of basophils rather than corticotrophs, which might be either thyrotrophs or gonadotrophs.
The immunohistochemical staining for PCNA showed that share of the immunoreactive cells was small in all the three age groups and it appeared to decrease with age.
Image analysis demonstrated that the volume density of the ACTH- immunoreactive cells did not change during preweaning period, remaining almost at the same level, with its mild reduction by the beginning of the infant period of life (p>0,05), while staining for PCNA showed significant decrease of the immunoreacive cells volume density (p<0,05) in the peripubertal age. This finding may be explained by the increased rate of differentiation of the corticotrophs with age during early prenatal development which compensated the relatively reduced proliferative potential of the adenocytes.
The results obtained provide evidence that in prepubertal rats the population of the corticotrophs in the adenohypophysis undergoes dynamic changes which predetermine age-related modulation in the hypothalamo-hypophyseo-adrenal axis in rats during early postnatal development and promotes better understanding of its activation potential in the growing body of experimental animals.
The work is submitted to the International scientific conference "Modern science technology", Tenerife, Spain, November, 20-27, 2008, came to the editorial office on 22.08.2008.