Nerve growth factor promoter activity revealed in mice expressing enhanced green fluorescent protein

Abstract

Nerve growth factor (NGF) and its precursor proNGF are perhaps the best described growth factors of the mammalian nervous system. There remains, however, a paucity of information regarding the precise cellular sites of proNGF/NGF synthesis. Here we report the generation of transgenic mice in which the NGF promoter controls the ectopic synthesis of enhanced green fluorescent protein (EGFP). These transgenic mice provide an unprecedented resolution of both neural cells (e.g., neocortical and hippocampal neurons) and non-neural cells (e.g., renal interstitial cells and thymic reticular cells) that display NGF promoter activity from postnatal development to adulthood. Moreover, the transgene is inducible by injury. At 2 days after sciatic nerve ligation, a robust population of EGFP-positive cells is seen in the proximal nerve stump. These transgenic mice offer novel insights into the cellular sites of NGF promoter activity and can be used as models for investigating the regulation of proNGF/NGF expression after injury. INDEXING TERMSneurotrophin; nerve growth factor; transgene; brain; peripheral nerve; injury Nerve growth factor (NGF), one of several neurotrophins of the vertebrate nervous system, was first localized in the male mouse submaxillary gland (Levi-Montalcini and Cohen, 1960). Later studies found that sympathetic ganglia, as well as target sites of sympathetic innervation (e.g., irides, heart atria, and prostate), have detectable levels of NGF mRNA and protein (Korsching and Thoenen, 1983a;Heumann et al., 1984;Shelton and Reichardt, 1984). The presence of NGF protein in sympathetic neurons has been linked to the binding and retrograde transport of this neurotrophin from target tissues, rather than to endogenous CIHR Author Manuscript CIHR Author Manuscript CIHR Author Manuscriptsynthesis by ganglionic neurons (or adjacent glia) (Hendry et al., 1974;Stöckel et al., 1976;Johnson et al., 1978).Intracerebral injections of iodinated NGF into selected sites, including the neocortex and hippocampus, revealed that the basal forebrain cholinergic neurons are capable of binding and retrogradely transporting NGF (Seiler and Schwab, 1984). This observation suggested that endogenous NGF might be utilized by these central projection neurons, in a manner similar to how NGF is acquired by sympathetic neurons from their peripheral target tissues. In the rat brain, levels of NGF mRNA are, in fact, highest in the neocortex and hippocampus, and select neuronal populations in both these central sites display NGF mRNA expression and protein synthesis (Korsching et al., 1985;Shelton and Reichardt, 1986;Whittemore et al., 1986;Ernfors et al., 1990;Lauterborn et al., 1991;. Detection of NGF mRNA by in situ hybridization and NGF protein by immunostaining elsewhere in the mammalian brain led to both complementary (e.g., in the basal forebrain) and conflicting (e.g., in the striatum) observations. Other methods are needed to provide further insight into the central sources of NGF synthesis (during normal development and as ...

Myenteric expression of nerve growth factor and the p75 neurotrophin receptor regulate axonal remodeling as a consequence of colonic inflammation in mice

“…We have reported the generation of transgenic mice that express EGFP under the control of the NGF promoter (Kawaja et al, 2011). Only one population of neurons was detected in the PNS of these mice: those found in the myenteric plexus of the intestinal tract, including the distal colon (that tissue which was the focus of this investigation) ( Fig.…”

“…Genotyping of NGFpr-EGFP mice used two primers that span the NGF promoter (5'-TCGGTGAGTC AGGCTTCTCT-3') and the EGFP (5'-TGAGTTTGGACAAACCACAAC-3') cDNAs (Kawaja et al, 2011). Genotyping of the NGF null mutant mice used three primers: 5'/WT: 5'CAGGCAGAACCGTACACA GA-3'; 3'/WT: 5'-CTGTCACTCGGGCAGCTATT-3'; and, 3'/Neo 5'-CCTTCTATCGCCTTCTTGACG-3'.…”

“…Four lines of mice were used: 1) NGFpr-EGFP mice that display EGFP expression under the control of murine NGF promoter (Kawaja et al, 2011), 2) the progeny of NGF +/-parent mice (B6.129S7-Ngf tm1Gne /J) (Crowley et al, 1994) 3) mice with a knock-out of the p75 neurotrophin receptor (Lee et al, 1992), and 4) the progeny of SMP-NGF/p75 +/-male mice and p75 +/-female mice, where SMP-NGF mice produce NGF under the control of the smooth muscle alpha actin promoter (SMP) (Petrie et al, 2013).…”

“…The precise cellular sources of NGF in normal and inflamed tissues, more often than not, have remained unresolved. To address this issue of cellular synthesis of NGF we generated lines of transgenic mice that express enhanced green fluorescent protein (EGFP) under the control of the NGF promoter (Kawaja et al, 2011). One surprising finding by Kawaja et al in 2011 was the presence of EGFP + neurons in the gastro-intestinal myenteric plexus.…”

“…To address this issue of cellular synthesis of NGF we generated lines of transgenic mice that express enhanced green fluorescent protein (EGFP) under the control of the NGF promoter (Kawaja et al, 2011). One surprising finding by Kawaja et al in 2011 was the presence of EGFP + neurons in the gastro-intestinal myenteric plexus. In this study we have sought to resolve the importance of these neurons that display NGF promoter activity in the normal and inflamed colons of mice.…”