Elsevier

Molecular Brain Research

Volume 122, Issue 2, 30 March 2004, Pages 116-125
Molecular Brain Research

Research report
Rescue of NGF-deficient mice I: transgenic expression of NGF in skin rescues mice lacking endogenous NGF

https://doi.org/10.1016/j.molbrainres.2003.12.004Get rights and content

Abstract

Mice lacking a functional NGF gene (ngf−/− mice) have less than one third of the normal complement of sensory neurons, few sympathetic postganglionic neurons and die shortly after birth. We report here that transgenic expression of NGF under control of the K14 keratin promoter can rescue some elements of the peripheral nervous system and restore normal growth and viability to ngf−/− mice. While hybrid transgenic-ngf−/− mice (ngfTKOs) displayed marginal rescue of trigeminal ganglion neurons, the percentage of CGRP-positive neurons was restored to normal. Restoration of CGRP-positive terminals in skin and spinal cord was also found and accompanied by recovery of behavioral responses to noxious stimuli. ngfTKO mice displayed a normal number of superior cervical ganglion neurons and recovery of sympathetic innervation of skin. These results demonstrate that substitution of a functional NGF locus by a transgene directing expression largely to skin can result in normal growth and viability. Thus, the most vital functions of NGF are not dependent on faithful recapitulation of the normal spatiotemporal pattern of gene expression.

Introduction

Nerve growth factor (NGF) is the founding member of the neurotrophin growth factor family and was isolated because of its ability to promote survival and axonal outgrowth in developing sensory and sympathetic neurons [23], [24], [25], [26]. The temporal and spatial expression pattern of NGF was the basis for development of the neurotrophic hypothesis, which proposes that target-derived growth factors modulate neuronal survival and innervation density [22], [35], [38]. Analysis of ngf−/− mice verified the intrinsic role of NGF as a target-derived survival factor by demonstrating loss of small diameter sensory neurons and sympathetic postganglionic neurons [9]. In contrast to mice that lack NGF, mice that overexpress NGF in the skin (NGF-OE) exhibit hypertrophy of both sensory and sympathetic nervous systems [1], [2], [12]. One line of NGF-OE mice demonstrated a two-fold increase in the number of trigeminal and dorsal root ganglion sensory neurons and a 2.5 fold increase in the number of sympathetic neurons in the superior cervical ganglia [1], [2], [21], [32]. In addition, preferential increases were found in unmyelinated and thinly myelinated fibers [13], [43], a finding consistent with the types of axons lost in ngf−/− mice.

Mutant mice that lack NGF typically die within the first postnatal week, greatly limiting study of the impact of NGF withdrawal on neuron maturation and function in peripheral and central systems [9]. A likely cause underlying the premature death of ngf−/− mice is the failure of these animals to properly feed due to disruption of the sensory nervous system. To test whether target-derived NGF can rescue these deficits in the peripheral nervous system (PNS), mice that lack a functional endogenous NGF gene were crossed with mice that overexpress NGF in the periphery driven by the human K14-keratin promoter [1], [45]. Using this approach, hybrid transgenic-knockout mice, or ngfTKO mice, were produced that showed normal growth and viability. Transgenic expression of NGF restored the number of sympathetic neurons to normal levels, whereas the number of sensory neurons was only marginally improved. An increase in the proportion of peptidergic sensory neuron profiles did occur, however, and was accompanied by restoration of peripheral and central projections. These anatomical changes were paralleled by the ability of ngfTKO mice to detect noxious stimuli. The restored sensory function and viability of ngfTKO mice points to the importance of peripheral targets in providing critical trophic influences in the development and function of the PNS.

Section snippets

Isolation of mouse lines

NgfTKO mice were isolated by mating previously isolated ngf+/− mice [9] with NGF-OE mice that express the K14-NGF transgene [1]. Hybrid heterozygous knockout mice (NGF-OE/NGF+/−) obtained in the first round of mating were then bred to isolate hybrid homozygous knockout-NGF-OE mice (NGF-OE/ngf−/−). Hybrid breeding used the 47-8 NGF-OE or medium expresser transgenic line (described in Ref. [1]). This mouse line was used to avoid systemic effects of NGF overexpression that may be seen in mice

Transgenic-NGF rescues viability of ngf−/− mice

The introduction of a transgenic source of NGF driven by the K14 keratin promoter significantly affected survival of ngf−/− mice (Table 1). While no non-transgenic ngf−/− mice were viable at the time of genotype identification, ngf−/− mice carrying the K14-NGF transgene (ngfTKO mice) were identified at roughly 2/3 the predicted frequency (Table 1). The frequency of other genotypes present at the time of identification was not markedly different from the predicted number of births, suggesting

ngfTKO mice exhibit sensitivity to noxious stimuli

Neonatal ngf−/− mice typically do not respond to tail pinch stimuli [9]. To determine whether transgenic-NGF could restore behavioral sensitivity, a tail pinch behavioral assay was conducted on ngfTKO mice. Mice heterozygous for the NGF null mutation (ngf+/−) responded strongly to tail pinch (Table 5). Mice heterozygous for both the NGF transgene and for the NGF null mutation (NGF-OE/ngf+/−) expressed strong responses to tail pinch in 15 of 17 cases. Similarly, 18 out of 20 ngfTKO mice

Discussion

Data from over half a century of research supports a role for NGF in development and function of selected subpopulations of neurons in both the central and peripheral nervous system. The biological importance of NGF is underscored by the observation that ngf−/− mice demonstrate loss of sympathetic and small diameter sensory neurons and typically die within the first postnatal week after exhibiting poor feeding and lack of growth [9]. While these findings confirm the importance of NGF in

Acknowledgements

This work was funded in part by the NIH from NS33730 (KMA), NS31826 (BMD), and by Genentech.

References (45)

  • J. Lu et al.

    Small primary sensory neurons innervating epidermis and viscera display differential phenotype in the adult rat

    Neurosci. Res.

    (2001)
  • B. Mendelson et al.

    Overexpression of nerve growth factor in epidermis of transgenic mice preserves excess sensory neurons but does not alter the somatotopic organization of cutaneous nerve projections

    Neurosci. Lett.

    (1996)
  • D.C. Molliver et al.

    IB4-binding DRG neurons switch from NGF to GDNF dependence in early postnatal life

    Neuron

    (1997)
  • T.D. Patel et al.

    Development of sensory neurons in the absence of NGF/TrkA signaling in vivo

    Neuron

    (2000)
  • F.L. Rice et al.

    Differential dependency of unmyelinated and A delta epidermal and upper dermal innervation on neurotrophins, trk receptors, and p75LNGFR

    Dev. Biol.

    (1998)
  • K.M. Albers et al.

    Overexpression of nerve growth factor in epidermis of transgenic mice causes hypertrophy of the peripheral nervous system

    J. Neurosci.

    (1994)
  • K.M. Albers et al.

    Overexpression of neurotrophin NT-3 in transgenic mice alters sensory receptor phenotype

    J. Cell Biol.

    (1996)
  • S. Averill et al.

    Immunocytochemical localization of trkA receptors in chemically identified subgroups of adult rat sensory neurons

    Eur. J. Neurosci.

    (1995)
  • V.L. Buchman et al.

    Different neurotrophins are expressed and act in a developmental sequence to promote the survival of embryonic sensory neurons

    Development

    (1993)
  • K. Chung et al.

    Sympathetic sprouting in the dorsal root ganglia of the injured peripheral nerve in a rat neuropathic pain model

    J. Comp. Neurol.

    (1996)
  • D. Creedon et al.

    Nerve growth factor synthesis in vascular smooth muscle

    Hypertension

    (1991)
  • A. Davies et al.

    Relation of target encounter and neuronal death to nerve growth factor responsiveness in the developing mouse trigeminal ganglion

    J. Comp. Neurol.

    (1984)
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