Distribution and Abundance of Glucocorticoid and Mineralocorticoid Receptors throughout the Brain of the Great Tit (Parus major)

Abstract

The glucocorticoid stress response, regulated by the hypothalamic-pituitary-adrenal (HPA) axis, enables individuals to cope with stressors through transcriptional effects in cells expressing the appropriate receptors. The two receptors that bind glucocorticoids-the mineralocorticoid receptor (MR) and glucocorticoid receptor (GR)-are present in a variety of vertebrate tissues, but their expression in the brain is especially important. Neural receptor patterns have the potential to integrate multiple behavioral and physiological traits simultaneously, including self-regulation of glucocorticoid secretion through negative feedback processes. In the present work, we quantified the expression of GR and MR mRNA throughout the brain of a female great tit (Parus major), creating a distribution map encompassing 48 regions. This map, the first of its kind for P. major, demonstrated a widespread but not ubiquitous distribution of both receptor types. In the paraventricular nucleus of the hypothalamus (PVN) and the hippocampus (HP)-the two brain regions that we sampled from a total of 25 birds, we found high GR mRNA expression in the former and, unexpectedly, low MR mRNA in the latter. We examined the covariation of MR and GR levels in these two regions and found a strong, positive relationship between MR in the PVN and MR in the HP and a similar trend for GR across these two regions. This correlation supports the idea that hormone pleiotropy may constrain an individual's behavioral and physiological phenotype. In the female song system, we found moderate GR in hyperstriatum ventrale, pars caudalis (HVC), and moderate MR in robust nucleus of the arcopallium (RA). Understanding intra- and interspecific patterns of glucocorticoid receptor expression can inform us about the behavioral processes (e.g. song learning) that may be sensitive to stress and stimulate future hypotheses concerning the relationships between receptor expression, circulating hormone concentrations and performance traits under selection, including behavior.

Fig 1. Image segmentation in Image J.

A custom macro segmented images of silver grains over cell bodies into separate cell body (A, top) and silver grain (A, bottom) binary images. Expression was calculated as the pixels of silver grains superimposed over cell bodies subtracting a background level of silver grains all divided by the cell body area (B). This allowed us to control for variation in the number of cell bodies present in different regions.

Fig 2. Plate 6 (A4.23–4.41 in the zebra finch atlas).

Legend (left) and map (right). GR (blue, left) and MR (red, right) expression is indicated by dot size. An “X” indicates that the background expression met or exceeded the expression in the region. Area abbreviations can be found in Table 2.

Fig 3. Plate 12 (~A2.97 in the zebra finch atlas).

Legend (left) and map (right). GR (blue, left) and MR (red, right) expression is indicated by dot size. An “X” indicates that the background expression met or exceeded the expression in the region. Area abbreviations can be found in Table 2.

Fig 4. Plate 17 (A2.07–2.25 in the zebra finch atlas).

Legend (left) and map (right). GR (blue, left) and MR (red, right) expression is indicated by dot size. An “X” indicates that the background expression met or exceeded the expression in the region. Area abbreviations can be found in Table 2.

Fig 5. Plate 21 (A1.53–1.71 in the zebra finch atlas).

Legend (left) and map (right). GR (blue, left) and MR (red, right) expression is indicated by dot size. An “X” indicates that the background expression met or exceeded the expression in the region. Area abbreviations can be found in Table 2.

Fig 6. Plate 28 (A0.36 in the zebra finch atlas).

Legend (left) and map (right). GR (blue, left) and MR (red, right) expression is indicated by dot size. An “X” indicates that the background expression met or exceeded the expression in the region. Area abbreviations can be found in Table 2.

Fig 7. Plate 30 (A0.18–0.00 in the zebra finch atlas).

Legend (left) and map (right). GR (blue, left) and MR (red, right) expression is indicated by dot size. An “X” indicates that the background expression met or exceeded the expression in the region. Area abbreviations can be found in Table 2.

Fig 8. Representative photopictomicrographs of MR and GR expression.

Cerebellum (Cb), paraventricular nucleus of the hypothalamus (PVN), hippocampus (HP), HVC, and nucleus robustus arcopallii (RA). Cell bodies are dark purple, silver grains are black dots. Scale bar indicates 50 μm.

Fig 9. Expression of GR (A) and MR (B) regressions across HP and PVN.

Units are area of silver grains superimposed over cell bodies minus background silver grain area divided by cell body area (see methods). Trendline and tests performed with log-transformed data, though untransformed data are shown. MR levels in HP significantly predicted MR levels in PVN (R² = 0.5941, p<0.0001). GR levels in HP demonstrated a trend toward predicting GR levels in the PVN (R² = 0.15, p = 0.069). No other combinations of areas and receptors were significant (all p>0.05).