Cellular signals underlying β-adrenergic receptor mediated salivary gland enlargement.

We examined the cellular signaling pathways involved in parotid gland enlargement induced by repeated isoproterenol administration in rats. Immunoblot analysis revealed early (1h) activation of the mitogen activated protein kinase (MAPK) ERK1/2, and progressive activation of epidermal growth factor receptor (EGFR), p38MAPK and p70S6 kinase (p70S6K) during 72h of isoproterenol treatment. Expression of β-adrenergic receptors (ARs) of the β2, but not β1, subtype increased over time in parallel with increases in the proliferation marker PCNA and parotid gland weight. Levels of β2-AR mRNA, assessed by quantitative RT-PCR and Northern blot analysis, were upregulated in parotid glands of isoproterenol treated rats. cAMP response element binding protein (CREB), a positive regulator of β2-AR transcription, was activated at 1h after isoproterenol administration, as evidenced by increased nuclear translocation and DNA binding using immunohistochemical staining and electrophoretic mobility shift assay. ELISA of NF-κB, also a β2-AR transcriptional regulator, revealed an increase in p65 and p50 subunits in nuclear protein extracts from parotid glands of isoproterenol treated rats. Together, these results demonstrate that β-adrenergic stimulation activates diverse cell survival and progrowth signaling pathways, including cAMP and EGFR linked activation of ERK1/2, p38MAPK, and p70S6K, and also induction of β2-ARs, possibly mediated by CREB and NF-κB, resulting in salivary gland enlargement. We propose that during isoproterenol treatment activation of the β1-AR, the predominant β-AR subtype in unstimulated salivary glands, initiates proliferative signaling cascades, and that upregulation of the β2-AR plays an essential role in later stages of salivary gland growth.