Mapping of CGRP in the alpaca (Lama pacos) brainstem

https://doi.org/10.1016/j.jchemneu.2008.02.004Get rights and content

Abstract

In this study, we demonstrate the presence of immunoreactive structures containing calcitonin gene-related peptide in the alpaca brainstem. This is the first time that a detailed mapping of the cell bodies and fibers containing this neuropeptide in the alpaca brainstem has been carried out using an immunocytochemical technique. Immunoreactive cell bodies and fibers were widely distributed throughout the alpaca brainstem. A high density of calcitonin gene-related peptide-immunoreactive perikarya was found in the superior colliculus, the dorsal nucleus of the raphe, the trochlear nucleus, the lateral division of the marginal nucleus of the brachium conjunctivum, the motor trigeminal nucleus, the facial nucleus, the pons reticular formation, the retrofacial nucleus, the rostral hypoglossal nucleus, and in the motor dorsal nucleus of the vagus, whereas a high density of fibers containing calcitonin gene-related peptide was observed in the lateral division of the marginal nucleus of the brachium conjunctivum, the parvocellular division of the alaminar spinal trigeminal nucleus, the external cuneate nucleus, the nucleus of the solitary tract, the laminar spinal trigeminal nucleus, and in the area postrema. This widespread distribution indicates that the neuropeptide studied might be involved in multiple functions in the alpaca brainstem.

Introduction

The family of calcitonin gene-related peptide and related peptides comprises amylin, adrenomedullin, calcitonin and calcitonin gene-related peptide. This latter peptide (composed of 37 amino acids) originates from the calcitonin gene. In studies carried out on mammals, mainly in the 1980s, fibers and cell bodies containing calcitonin gene-related peptide were observed to be widely distributed thought the mammalian central nervous system (Skofitsch and Jacobowitz, 1985a; see Palkovits, 1988, Batten et al., 1989, Conti and Sternini, 1989, McWilliam et al., 1989, Tashiro et al., 1991, Unger and Lange, 1991, Quartu et al., 1992, Marcos et al., 1999, Coveñas et al., 2003a), and by radioimmunoassay, the quantitative distribution of calcitonin gene-related peptide was reported in the rat central nervous system (Skofitsch and Jacobowitz, 1985b). Moreover, also since the 1980s, the distribution of the calcitonin gene-related peptide binding sites in the mammalian central nervous system has been known (Skofitsch and Jacobowitz, 1985c, Tschopp et al., 1985).

The calcitonin gene-related peptide has been implicated in the regulation of the differentiation and activation of astrocytes, in anorexia, inflammation, analgesia, hyperthermia, catalepsia, and the regeneration of peripheral nerves. It also inhibits gastric acid secretion, relaxes the muscle cells of the urinary tract, produces vasodilatation, and inhibits acid secretion (HCl) by oxyntic cells, inhibiting the formation of ulcers in the intestinal or gastric mucosae. It also decreases locomotor activity, controls the expression of neuropeptides/neurotransmitters by neurons, regulates social behavior, inhibits peristalsis and enzymatic secretion by pancreatic acinar cells and the secretion of insulin by beta cells, and regulates hearing (Maison et al., 2003a, Maison et al., 2003b; see Coveñas et al., 2007).

In general, in South America camelids previous research has been mainly focused on the reproductive mechanisms of these animals (Bravo et al., 1996, Correa et al., 1997, Ratto et al., 1997, Ratto et al., 2005, Ratto et al., 2006), due to the economic importance that the wool of such animals has in some South America countries. The authors of these works focused their attention on the hormonal indicators of pregnancy in llamas and alpacas, on superovulation in llamas caused by the administration of follicle stimulating hormone, and on the effects of hormonal treatments and natural mating on ovulation induction, interval to ovulation, and luteal development in llamas.

However, no previous information appears to be available in the literature concerning the presence of fibers and cell bodies containing neuropeptides in the alpaca or llama central nervous systems following the implementation of immunocytochemical techniques, except for a recent study carried out by our group on the distribution of immunoreactive structures containing leucine–enkephalin in the alpaca brainstem (de Souza et al., 2007). In this sense, the chemical neuroanatomy of the neuropeptides in the alpaca brainstem has not yet been studied in depth, a field that must be developed in the future in order to better understand the physiology (e.g., the involvement of neuropeptides at altitude, reproductive behavior, etc.) of this camelid. In light of the above, the main aim in this work was to study for the first time the distribution of fibers and cell bodies containing calcitonin gene-related peptide in the alpaca (Lama pacos) mesencephalon, pons and medulla oblongata, using an immunocytochemical technique. Another aim was to compare the distribution of calcitonin gene-related peptide-immunoreactive structures in the alpaca brainstem with that of those located in the same region of the alpaca central nervous system containing leucine–enkephalin (de Souza et al., 2007).

Section snippets

Animals

Five male adult alpacas (L. pacos) (70–80 kg) were obtained from the Peruvian University Cayetano Heredia (Faculty of Veterinary Medicine and Animal Sciences, Lima, Peru). The animals were kept (altitude: 0 m) under standard conditions of light and temperature and had free access to food and water. All the animals, used in this study, were always maintained at 0 m on the sea level (from the birth to the perfusion).

The experimental design, protocols, and procedures of this work were performed under

General considerations

The presence of immunoreactive structures containing calcitonin gene-related (CGRP) peptide in the alpaca brainstem is shown in Fig. 1 and Table 1. These show a widespread distribution of both CGRP-immunoreactive fibers and cell bodies. We found numerous immunoreactive cell bodies distributed throughout the whole alpaca brainstem (see Fig. 1), although the four animals studied here were not treated with colchicine. In general, in all the animals studied the distribution and density of both

CGRP in the alpaca brainstem

This is the first time that the presence of fibers and cell bodies containing CGRP in the alpaca central nervous system has been reported using an immunocytochemical technique. The distribution of the immunoreactive structures (fibers and cell bodies) containing CGRP was widespread in the alpaca brainstem, although it should be noted that the animals used in this study were not treated with colchicine. Despite this, we found a high number of immunoreactive cell bodies containing CGRP. This

Acknowledgements

This work has been supported by the Ministerio de Educación y Ciencia (BFU2005-02241/BFI), Spain and by the CONCYTEC: PROCYT project 2006, Peru. The authors thank Ewing Rafael Duque for his technical assistance, N. Skinner for stylistic revision of the English text and Professor Gerard Tramu (University of Bordeaux I, France) for kindly providing the CGRP antiserum.

References (43)

  • A. Mangas et al.

    Immunocytochemical visualization of d-glutamate in the rat brain

    Neuroscience

    (2007)
  • P. Marcos et al.

    Immunohistochemical mapping of enkephalins, NPY, CGRP and GRP in the cat amygdala

    Peptides

    (1999)
  • I. Marshall et al.

    Cardiovascular effects of human and rat CGRP compared in the rat and other species

    Eur. J. Pharmacol.

    (1986)
  • P.K. Mulderry et al.

    Distribution and chromatographic characterization of CGRP-like immunoreactivity in the brain and gut of the rat

    Regul. Pept.

    (1985)
  • P. Pesini et al.

    Distribution of ACTH immunoreactivity in the diencephalons and brainstem of the dog

    J. Chem. Neuroanat.

    (2004)
  • M. Quartu et al.

    Calcitonin gene-related peptide in the human trigeminal sensory system at developmental and adult life stages: immunohistochemistry, neuronal morphometry and coexistence with substance P

    J. Chem. Neuroanat.

    (1992)
  • M.H. Ratto et al.

    Timing of mating and ovarian response in llamas (Lama glama) treated with pFSH

    Anim. Reprod. Sci.

    (1997)
  • M.H. Ratto et al.

    Comparison of the effect of natural mating, LH, and GnRH on interval to ovulation and luteal function in llamas

    Anim. Reprod. Sci.

    (2006)
  • G. Skofitsch et al.

    Calcitonin gene-related peptide: detailed immunohistochemical distribution in the central nervous system

    Peptides

    (1985)
  • G. Skofitsch et al.

    Quantitative distribution of calcitonin gene-related peptide in the rat central nervous system

    Peptides

    (1985)
  • G. Skofitsch et al.

    Autoradiographic distribution of 125I calcitonin gene-related peptide-binding sites in the rat central nervous system

    Peptides

    (1985)
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