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Original Paper


              The results of this study have allowed the       2004). In many teleost fish studied, immunohisto-
             immunolocalization of the distribution of GnRH-   chemistry highlighted sGnRH immunoreactivity of
             like structures in the brain and pituitary gland of  the neurons of the anterior ventral brain (telen-
             bluefin tuna.                                     cephalon and preoptic region), whereas the large
              Eight GnRH variants have been identified in the  cell bodies of the midbrain tegmentum were found
             brain of teleost fish,but (c) GnRH II,(s) GnRH and  to react to anti cGnRH-II serum (see Lethimonier
             (sb) GnRH have been found to coexist in some per-  et al., 2004).
             ciformes and other fishes (Lethimonier  et al.,    The perikarya of the preopticus-periventricularis
             2004).                                            nucleus immunoreacted with anti sGnRH 1667,
              Four different polyclonal antibodies against     anti cGnRH-II 675 and anti cGnRH-II 6 sera.
             sGnRH, cGnRH and sbGnRH were used. In this        Differences in the distribution, % and size of the
             study three antisera (sGnRH 1667, cGnRH-II 675,   immunoreactive perikarya were observed. Anti
             cGnRH-II 6) revealed immunoreactivity in the      sGnRH- and anti cGnRH-positive cells were mainly
             perikarya of the oculomotor nucleus, anterior mid-  found in the preopticus and caudal part of periven-
             brain tegmentum, preopticus-periventricular and   tricularis nucleus. sGnRH-immunoreactive cells
             lateralis tuberis nuclei in the diencephalon, the neu-  comprised around 20-30% of the total whereas
             rohypophysis and in the junction between olfactory  cGnRH-like neurons comprised around 50%. Anti
             bulbs and telencephalon.                          sGnRH-immunoreactive neurons were smaller than
              Surprisingly no immunoreactive structures were   the anti cGnRH ones. The presence of GnRH neu-
             found with anti sbGnRH serum although sbGnRH is   rons in the preopticus nucleus has been detected as
             thought to be the most important regulator of     a sGnRH variant in Cypryniformes (Yu et al., 1988;
             gonadal function (Lethimonier et al., 2004; Pham  Powel et al., 1996) and Salmoniformes (Okuzawa
             et al., 2006), and sbGnRH-producing cell bodies   et al., 1990; Amano  et al., 1991), whereas the
             project their axons mainly to the pituitary gland  cGnRH-II form has been reported in midbrain
             (White  et al., 1995; Gonzalez-Martinez  et al.,  tegmentum of most of the fish species studied (see
             2001).This negative result is of difficult interpreta-  Lethimonier  et al., 2004 for review). In bluefin
             tion despite the fact that different dilutions and  tuna, we were able to find weakly immunostained
             incubation times were used in the experiments car-  nerve fibres in both the preopticus nucleus and the
             ried out here. The lack of anti sbGnRH immunos-   neurohypophysis.This is in line with the function of
             taining likely depends on technical reasons, includ-  the preopticus nucleus which is considered to be the
             ing the non-specificity of the primary antibody   main source of the innervation of the neurohypoph-
             used, as well as likely antigen damage during the  ysis (Yamamoto et al., 1998).
             fixation and paraffin wax embedding. However,      The nucleus the lateralis tuberis exhibited
             since large differences in the levels of sbGnRH have  immunostaining with anti sGnRH 1667 serum in
             been shown among perciform  fishes (Senthil-      40% of the neurons, whereas it did not react with
             kumaran  et al., 1999), a possible explanation is  antisera against cGnRH and sbGnRH. The occur-
             that the bluefin tuna contains such a low amount of  rence of GnRH cells in this nucleus has been
             the sbGnRH form that it becomes undetectable to   observed in catfish  Clarias batrachus (Subhedar
             the immunostaining.Therefore, other types of stud-  and Rama Krishna, 1988; Khan  et al., 1999;
             ies,including RIA and in situ hybridization might be  Sarkar and Subhedar, 2000). GnRH cells have been
             helpful in finding a solution to this still unanswered  reported in the caudo-distal pars of neonatal platy-
             question.                                         fish (Schreibman et al., 1982).
              cGnRH-II is expressed in all teleosts examined,   In this study, we were able to find neurons and
             including sarcopterigy and actinopterigy (review-  nerve fibres immunoreactive to cGnRH and sGnRH
             ered by Letimonier et al., 2004). With the excep-  sera in the bulbs as well as in the olfactory nerve of
             tions of the butterfly fish (O’neill et al., 1998), the  bluefin tuna. According to several authors, the
             European eel (King  et al., 1990) and the catfish  GnRH-immunoreactive cells detected in this tract
             (Ngamvongchon  et al., 1992), sGnRH has been      of the brain are the cells most strongly subjected to
             detected in all teleost species from primitive fish,  the action of pheromones (Fujita  et al., 1991;
             osteoglossiformes (Okubo and Aida, 2001; O’neill  Egorova et al., 2001) and are likely to be involved
             et al., 1998), to acanthopterigii (Lethimonier et al.,  in fish sexual behaviour neuromodulation (Oka and

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