Localization of Calbindin-D28k and Calretinin in the Brain of Dermophis Mexicanus (Amphibia: Gymnophiona) and Its Bearing on the Interpretation of Newly Recognized Neuroanatomical Regions

Abstract

The analysis of the distribution of the calbindin-D28k and calretinin immunoreactive (CBir and CRir) systems recently described in the brain of anuran and urodele amphibians was very useful for the interpretation of many otherwise indistinct brain regions and cell masses. In the present study we have followed a similar approach to investigate the distribution of CBir and CRir cell bodies and fibers in the brain of Dermophis mexicanus, a member of the much neglected third amphibian order of gymnophionans. The pattern of distribution obtained showed particular characteristics in Dermophis, such as the existence of abundant CRir elements in the olfactory bulbs and CBir and CRir cell populations in pallial areas. The distinct distribution of the two proteins allowed the tentative identification of currently described subregions, mainly in the amygdaloid complex and hypothalamic areas. The analysis of the diencephalon and brainstem distribution framed in the neuromeric model highlighted common traits with other amphibians but also specific features. Therefore, the immunohistochemical detection of calcium-binding proteins has served to discern cell populations and has helped to demonstrate neuronal heterogeneity. However, it should be pointed out that a straightforward comparison based only on the presence of these proteins should not be made due to the great variability observed in well-established homologous regions in the brain of different vertebrates, as evidenced within the class Amphibia.

Immunohistochemical distribution of calretinin and calbindin (D‐28k) in the brain of the cladistian Polypterus senegalus

“…As regards the presence of CB-ir populations, CB-ir thalamic cells have been reported in a dogfish (Rodríguez-Moldes et al, 1990a), the Siberian sturgeon (Graña et al, 2012) and the Senegal bichir (present results). All amphibians investigated showed CB-ir cell populations in the thalamus (Morona and Gonz alez, 2008;Morona et al, 2011a).…”

“…Both proteins act as intracellular calcium buffers (Miller, 1995), neuromodulators (Schwaller et al, 2002), or calcium sensors (McCue et al, 2010), among other functions. Antibodies raised against these proteins have been widely used as neuronal markers to characterize neuronal populations or brain regions in tetrapods (mammals: García-Segura et al, 1984;Celio, 1990;S equier et al, 1990;Jacobowitz and Winsky, 1991;Resibois and Rogers, 1992;Andressen et al, 1993;birds: Roth et al, 1981;Rogers et al, 1990;Puelles et al, 1994;reptiles: D avila et al, 1997amphibians: Necchi et al, 1999;Mil an and Puelles, 2000;Gonz alez, 2008, 2009;Morona et al, 2011a) and fishes (lampreys: Pombal and Puelles, 1999;Megías et al, 2003;elasmobranchs: Rodríguez-Moldes et al, 1990a,b;Anad on et al, 2009;chondrosteans: Graña et al, 2012;holosteans: Northcutt, 2009;teleosts: Díaz-Regueira and Anad on, 2000;Castro et al, 2003Castro et al, , 2006aNorthcutt, 2009;lungfish: Gonz alez et al, 2010;Morona et al, 2010).…”

Regional chemoarchitecture of the brain of lungfishes based on calbindin D‐28K and calretinin immunohistochemistry

“…Thus, these proteins have served as anatomical markers in numerous studies in all classes of tetrapods (birds: Roth, Baetens, Norman, & García‐Segura, ; Rogers et al, ; Puelles, Robles, Martínez‐de‐la‐Torre, & Martínez, ; Belekhova et al, ; reptiles: Dávila, Padial, Andreu, Real, & Guirado, ; Dávila, Guirado, & Puelles, ; Yan, Tang, & Carr, ; Song, Wang, Carr, Dai, & Tang, ; amphibians: Necchi, Soldani, Pisu, Bernocchi, & Scherini, ; Milán & Puelles, ; Morona & González, ; Morona, López, & González, ). In particular, analysis of the distribution of CB‐ and CR‐containing cells and fibers has been widely conducted in the brain of many fish groups to assist in the identification of regions and nuclei across species because their staining, in many cases, facilitates topological recognition of homologous structures (lampreys: Pombal & Puelles, ; Megías, Álvarez‐Otero, & Pombal, ; elasmobranchs: Rodríguez‐Moldes et al, ; Rodríguez‐Moldes et al, ; Anadón et al, ; all groups of actinopterygians: Díaz‐Regueira & Anadón, ; Castro, Becerra, Manso, & Anadón, ,b; Northcutt, ; Jadhao & Malz, ; Graña, Huesa, Anadón, & Yáñez, ; Graña, Folgueira, Huesa, Anadón, & Yáñez, ; Deshpande & Jadhao, ).…”

“…Previous studies in the different groups of amphibians have demonstrated that the immunohistochemical localization of CB and CR is a powerful method for revealing discrete neuronal populations, fiber tracts, and neuropils (Milán & Puelles, ; Morona & González, ; Morona et al, ). Moreover, the patterns observed were consistent with the conceptual subdivisions of the segmental paradigm of the brain, and the analysis of the results in that framework enabled comparisons across species of different vertebrate classes (Pombal & Puelles, ; Dávila, Guirado, & Puelles, ; González, Puelles, & Medina, ; Puelles & Rubenstein, ).…”

“…We have used double‐labeling techniques for the simultaneous observation of CB and CR. Most previous investigations were restricted to the single immunolabeling of one protein, although their combined staining would provide more information on relationships among neurons containing different calcium‐binding proteins (Morona, López, & González, ,b; Morona, López, Domínguez, & González, ; Morona, Moreno, López, & González, ; Morona, López, & González, ; Morona, Northcutt, & González, ; Morona & González, ). We found that these two proteins are specifically expressed in distinct neuronal groups, helping to characterize cytoarchitectonically indistinguishable regions.…”