Expression of mRNAs for neuropeptide receptors and β-adrenergic receptors in human osteoblasts and human osteogenic sarcoma cells

Understanding of the mechanisms of neuropeptide gene regulation has increased dramatically in the past 5 years. In large part, this increased understanding has resulted from advances in the technology of introducing reporter genes into intact cells in culture. This chapter addresses two methods for gene transfer that have been particularly important for studies of neuropeptide gene regulation:DNA-mediated gene transfer into mammalian cell lines and microinjection of foreign genes into the germline of transgenic mice. Two specific areas of gene regulation will be considered — transcriptional control by cAMP and tissue-specific expression of neuropeptides in the brain and pituitary gland.

Regulation of neuropeptide gene expression

The ENS is responsible for the regulation and control of all gastrointestinal functions. Because of this critical role, and probably as a consequence of its remarkable plasticity, the ENS is often relatively well preserved in conditions where the architecture of the intestine is seriously disrupted, such as in IBD. There are structural and functional changes in the enteric innervation in animal models of experimental intestinal inflammation and in IBD. These include both up and down regulation of transmitter expression and the induction of new genes in enteric neurons. Using Fos expression as a surrogate marker of neuronal activation it is now well established that enteric neurons (and also enteric glia) respond to inflammation. Whether this "activation" is limited to a short-term functional response, such as increased neuronal excitability, or reflects a long-term change in some aspect of the neuronal phenotype (or both) has yet to be firmly established, but it appears that enteric neurons are highly plastic in their response to inflammation.

https://anatomypubs.onlinelibrary.wiley.com/doi/pdf/10.1002/1097-0185%2820010101%29262%3A1%3C79%3A%3AAID-AR1013%3E3.0.CO%3B2-K

Consequences of intestinal inflammation on the enteric nervous system: Neuronal activation induced by inflammatory mediators

Cocaine- and amphetamine-regulated tran- script (CART) peptide, originally isolated from brain, is also expressed in the peripheral nervous system. The distribution,originandprojectionsofCART-expressing enteric neurones by immunocytochemistry and in situ hybridization in rat gastrointestinal (GI) tract were studied. Possible motor functions of CART were stud- ied in vitro using longitudinal muscle strips from stomach,ileumandcolon.Cocaine-andamphetamine- regulated transcript peptide was found in numerous myenteric neurones throughout the GI tract while CART-expressing submucous neurones were scarce. Cocaine- and amphetamine-regulated transcript was also expressed in the antral gastrin cells. Myenteric CART-expressing neurones in both small and large intestine issued short descending projections. In atrophic ileum, CART mRNA-expressing neurones increased in number while neurones containing CART peptidedecreased.Inhypertrophiedileum,nochangein CART peptide or CART mRNA containing myenteric neurones was detected. Cocaine- and amphetamine- regulatedtranscript55-102(10 )9 -10 )7 mol L )1 )didnot induce any contractile or relaxatory responses in the muscle strips, neither did it affect responses induced by vasoactive intestinal peptide, pituitary adenylate cyclase-activating peptide or neuronal stimulation. In colonic, but not in ileal, strips addition of CART attenuated nitric oxide (NO) donor-induced relaxa- tions. Although CART does not seem to play a pivotal role in classic neurotransmission to the longitudinal muscle, it may serve a modulatory role in NO trans- mission. It may, moreover, be involved in intestinal adaptation, and an additional hormonal role is also

/pdf/cocaine-and-amphetamine-regulated-transcript-distribution-316w3r0jf5.pdf

Cocaine- and amphetamine-regulated transcript: distribution and function in rat gastrointestinal tract.

Several motility disorders originate in the enteric nervous system (ENS). Our knowledge of factors governing survival of the ENS is poor. Changes in the expression of vasoactive intestinal peptide (VIP) and nitric oxide synthase (NOS) in enteric neurons occur after neuronal injury and in intestinal adaptation. The aim of this study was to evaluate whether VIP and nitric oxide (NO) influence survival of cultured, dissociated myenteric neurons. Neuronal survival was evaluated after 0, 4, and 8 days in culture. Influence of VIP and NO on neuronal survival was examined after culturing in the presence of VIP, NO donor, VIP antiserum, or NOS inhibitor. A marked loss of neurons was noted during culturing. VIP and NO significantly promoted neuronal survival. Corroborating this was the finding of an enhanced neuronal cell loss when cultures were grown in the presence of VIP antiserum or NOS inhibitor.

Vasoactive intestinal peptide and nitric oxide promote survival of adult rat myenteric neurons in culture.

Expression of mRNA for vasoactive intestinal peptide in rat small intestine.

Gene expression and transcript size of the prepro-peptide VIP/PHM-27 in normal human tissue.

The availability of colon provides a ready source of human neurons. Among the products of nerve cell bodies, vasoactive intestinal peptide is a neuropeptide that serves as a marker of non-adrenergic, non-cholinergic inhibitory nerves in colon. These nerves have been proposed to be involved in regulation of immune function, secretion, and smooth muscle function. In previous work, we identified decreased tissue levels of vasoactive intestinal peptide in a disorder of chronic colonic mucosal inflammation, ulcerative colitis. We hypothesized that diminished gene expression of vasoactive intestinal peptide could result in decreased tissue levels of this neuropeptide. Sigmoid colon was obtained at surgery from controls (n = 6) and patients with ulcerative colitis (n = 6). Vasoactive intestinal peptide mRNA was quantified by Northern blot hybridization and tissue levels of vasoactive intestinal peptide were determined by radioimmunoassay. Tissue vasoactive intestinal peptide was decreased only in the mucosal-submucosal layer of ulcerative colitis (p = .02). There was a single 1.7 kbase vasoactive intestinal peptide transcript identified in both control colon and ulcerative colitis. Normalized vasoactive intestinal peptide mRNA levels were increased by 260% in ulcerative colitis compared to controls (p < .01). These observations suggest that decreased vasoactive intestinal peptide gene expression or abnormal post-transcriptional processing are not primary defects in this disorder of chronic inflammation. The findings support the alternative hypothesis that axonal degeneration in ulcerative colitis could result in increased expression of neuronal vasoactive intestinal peptide mRNA.

Expression of mRNA for vasoactive intestinal peptide in normal human colon and during inflammation.

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Expression of mRNA for vasoactive intestinal peptide in rat small intestine.

Gene expression and transcript size of the prepro-peptide VIP/PHM-27 in normal human tissue.

Expression of mRNA for vasoactive intestinal peptide in normal human colon and during inflammation.