Effects of chemical and surgical sympathectomy on expression of beta-adrenergic receptors and guanine nucleotide-binding proteins in rat submandibular glands

Abstract

Expression of beta 1-adrenergic receptors and guanine nucleotide-binding proteins in rat submandibular glands was determined after reserpine administration and sympathetic denervation. Pretreatment of rats with reserpine resulted in up-regulation of the density of beta 1-adrenergic receptors and the immunoreactivity of the 64-kDa species of beta 1-adrenergic receptor in submandibular membranes, by 2.6 +/- 0.3-fold (eight experiments), within 7 days. Steady state levels of beta 1-adrenergic receptor mRNA quantified by DNA-excess solution hybridization were 0.15 +/- 0.03 amol of beta 1-adrenergic receptor mRNA/micrograms of total cellular RNA (six experiments). beta 1-Adrenergic receptor mRNA increased by 50% within 8 hr after pretreatment with reserpine. Maximal levels of 0.37 +/- 0.04 amol of beta 1-adrenergic receptor mRNA/micrograms of RNA were attained by 4 days and these levels were sustained for the next 3 days (six experiments). Northern blot hybridization also revealed a 3-fold increase in the 2.5-kilobase beta 1-adrenergic receptor mRNA transcript, which was equivalent in magnitude to that determined by solution hybridization. Reserpine pretreatment also affected steady state levels of submandibular guanine nucleotide-binding proteins. Two immunoreactive forms of the alpha subunit of Gs, migrating as 42 kDa (major) and 50 kDa (minor), were detected in salivary membranes. The immunoreactivity of the 42-kDa species of Gs alpha declined by 50% after 7 days of continuous daily injections of reserpine. In contrast, steady state levels of Gi alpha 2 (41 kDa), Go (39 kDa), and G beta 2 (35 kDa) and their mRNAs in submandibular membranes were unaffected by reserpine pretreatment. The rate of beta 1-adrenergic receptor gene transcription assessed by nuclear run-on transcription assay in nuclei of submandibular glands was not altered by reserpine pretreatment. However, reserpine had a dramatic effect on the half-life of beta 1-adrenergic receptor mRNA in submandibular glands. The half-life of beta 1-adrenergic receptor mRNA in control submandibular glands was 3.5 hr, whereas it increased to 8 hr in reserpine-pretreated glands. Reserpine-promoted stabilization of beta 1-adrenergic receptor mRNA provides a mechanism for up-regulation of postjunctional beta 1-adrenergic receptors in sympathetically innervated tissues.