Elsevier

Brain Research

Volume 1041, Issue 2, 18 April 2005, Pages 157-166
Brain Research

Research report
Neurturin, persephin, and artemin in the human pre- and full-term newborn and adult hippocampus and fascia dentata

https://doi.org/10.1016/j.brainres.2005.02.007Get rights and content

Abstract

The immunochemical occurrence and localization of the Glial cell line-derived neurotrophic factor (GDNF) family ligands neurturin (NTN), persephin (PSP), and artemin (ART) is described in the human postmortem hippocampus and fascia dentata from subjects aged 21 weeks of gestation to 88 years. The detectability of NTN, PSP, and ART is shown in the rat by Western blot and immunohistochemistry up to 70 h postmortem. In the human tissue, labeled neuronal perikarya were detectable for each trophin at all examined ages, with prevalent localization in the pyramidal layer of the Ammon's horn and hilus and granular layer of the fascia dentata. In the adult subjects, punctate elements were also present. Comparison of the pattern of immunoreactive structures among young and adult subjects suggests that intracellular distribution and/or trafficking of the GDNF family ligands may undergo age-related changes. Labeled glial elements were also identifiable. Western blot analysis indicates that the availability of the dimeric and monomeric forms of the trophins may vary with age and postmortem delay. The results obtained suggest the involvement of NTN, PSP, and ART in processes subserving both the organization of this cortical region during development and the functional activity and maintenance of the mature human hippocampal neurons.

Introduction

Neurturin (NTN) [16], persephin (PSP) [24], and artemin/neublastin (ART) [4], [30], distant members of the transforming growth factor-β superfamily, are structurally related trophic proteins belonging to the glial cell line-derived neurotrophic factor (GDNF) family. They exert their biological actions via retrograde signalling through a multi-component tyrosine kinase receptor system [1], [35]. Expression analysis indicates that they occur in fetal and adult brain [4], [13], [16], [24] and behave as potent regulators of the development, differentiation and maintenance of mature phenotype for a wide variety of neuronal populations [4], [12], [13], [16], [24]. Further experimental evidence shows that they exert neuroprotective and neuroreparative activities in experimental brain injury and in animal models of neurodegeneration [11], [14], [15], [21], [22], [27], [29], [30], [34], [37], suggesting for them roles in neuropathogenesis and potential in clinical application. Studies on the expression of NTN in the hippocampus of experimental animals propose the involvement of this molecule in the development [12], [17], maintenance, and plasticity of hippocampal neuronal circuits [10], [15], [17]. Quantitative data indicate that levels of NTN mRNA are transiently high at embryonic and perinatal ages and become relatively low in the adult rat hippocampus [12], [17]. On the other hand, they are upregulated in the adult following insults such as epileptic seizures or forebrain ischemia [15]. Data on knockout mice suggest that NTN may modulate the seizure susceptibility of hippocampal neurons [26] and functional studies show the efficacy of ART in protecting hippocampal neurons against NMDA-induced excitotoxicity [6]. The potential relevance of NTN, PSP, and ART in the normal activity and in the pathological events involving the hippocampus underline the importance of gathering information concerning the involvement of these molecules in the trophism of the human hippocampal formation. As a first step in this line, this investigation analyses the immunohistochemical occurrence of NTN, PSP, and ART in the human archicortex. The possibility that the functional significance of NTN, PSP, and ART vary with age, as shown in the hippocampus and other neuronal systems in experimental animals [2], [4], [12], [17], [24], prompted us to extend the analysis to the widest range of ages we could examine, from prenatal life to adulthood.

Section snippets

Materials and methods

Autopsy specimens of human hippocampal formation were obtained from subjects of different ages, with no history of neuropathology (Table 1). The sampling and handling of human specimens has been conducted under the appropriate ethical safeguards and protocols of the Section of Pathological Anatomy, Department of Cytomorphology, University of Cagliari. Sixteen male adult Sprague–Dawley rats were sacrificed by decapitation and their head was kept at 4 °C for 0, 24, 48, and 70 h postmortem

Western blot

As shown in Fig. 1A, when challenged with the three trophins the anti-NTN and anti-ART sera labeled a single band at a level corresponding to the molecular weight (mw) of the monomeric form of the relevant trophin. In contrast, the anti-PSP serum showed two bands in the lane of the rhPSP, one at a mw corresponding to the monomeric form and a lighter one at a mw corresponding to the dimeric form of the protein. In addition, it showed a thick band in the rhART lane at the level of the ART

Western blot analysis

It has been reported that the monomeric form of the mature mouse and human NTN has a predicted molecular weight (mw) of 11.5 kDa [16]; however, other longer model sequences have been described (NCBI accession No.XP_217308) [33]. Thus, the band with a mw slightly higher than 25 kDa observed in the human tissue is consistent with the homodimeric, functionally active form of the protein [16]. As for PSP and ART, the bands observed in human tissue homogenates are consistent with the homodimer of

Acknowledgments

This work was funded by the Italian Consiglio Nazionale delle Ricerche (M.D.F.) and Fondazione Banco di Sardegna (M.Q.).

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