Cytokine regulation of neuropeptide expression in cultured sympathetic neurons

Abstract

This thesis discusses the regulation of substance P (SP) in cultured sympathetic neurons by different cytokines and compares SP to neuropeptide Y (NPY). The chapters are comprised of manuscripts submitted for publication or already in press.;Chapter one describes the regulation of SP in cultures of neonatal rat superior cervical ganglia (SCG) by interleukin-1{dollar}\beta{dollar} (IL-1{dollar}\beta{dollar}). IL-1{dollar}\beta{dollar} increased SP expression in neurons cocultured with ganglion nonneuronal cells, but not in pure neuronal cultures. Unlike other known mediators of SP, IL-1{dollar}\beta{dollar} did not effect cholinergic traits. As IL-1{dollar}\beta{dollar} did not stimulate SP in pure neuronal cultures, the cytokine probably acted on the Schwann cells which, in turn, activated SP expression in the neurons. Chapter two examines whether endogenous IL-1{dollar}\beta{dollar} regulates SP expression in the SCG. Incubation of cultured SCG's with an antagonist to the IL-1 receptor suppressed SP levels to 50% of control levels, suggesting that endogenous IL-1{dollar}\beta{dollar} at least partially regulates SP expression by these cultures. Western and Northern blot analyses of SCG neurons demonstrated that cultured sympathetic neurons synthesize and release IL-1{dollar}\beta{dollar}. The third chapter discusses SP regulation by another cytokine, leukemia inhibitory factor (LIF). LIF is a cholinergic and SP stimulating factor in both pure neuronal and in cocultures, distinguishing it from IL-1{dollar}\beta{dollar}. Depolarizing stimuli and dexamethasone treatment inhibited SP expression in SCG cultures, whereas LIF increased SP under either condition. Treatment of SCG cocultures with a monoclonal antibody directed against rat LIF partially blocked the development of cholinergic traits and SP, suggesting endogenous LIF also regulates the expression of transmitter properties. Chapter four describes the regulation of NPY, which, unlike SP, is expressed in 60% of SCG neurons. Neuronal density had a striking effect on NPY expression in both pure neuronal and cocultures; such that increased neuron number decreases neuronal NPY content. Moreover, levels of NPY were 2-fold greater in pure neuronal than in cocultures. LIF treatment decreased NPY slightly in pure neuronal cultures, but significantly in cocultures. Treatment with other immunoregulatory agents had no effect on the peptide. These observations indicate that NPY is regulated differently than SP in cultured sympathetic neurons.