**2.3 Microcrystalline silica in electric organs**

Based on the evidence of aluminum and silicium accumulation in *P. extenta,* we documented the presence of silica minerals in *P. extenta* electric tissue by means of mineralogical techniques (Prado Figueroa *et al*., 2008). It was thought that these compounds could form minerals (i.e., solid inorganic substances with a defined chemical composition and determined crystallography).

Fractionation of electric tissue homogenates by differential centrifugation was carried out as described for other tissues (Beaufay and Amar-Costesec, 1976; Amar-Costesec *et al*., 1985)

Fig. 2. Photomicrograph of a cytoplasmic extract of the electric organ from *P. extenta* in crossed-polarizers, by using a standard polarized light microscope.

The crossed-polarizer image of a cytoplasmic extract shows SiO2 replacements in grey and white arrangements, with undulatory extinction.

An electric organ cytoplasmic extract (50 μl) on lyophilised paper (Labconco Corp., USA) was fixed in 2.5% glutaraldehyde in a 0.05 M sodium phosphate buffer (pH 7.2) for 60 min at 4 °C. Samples were washed with buffer and bi-distilled water for 2 h, then, dehydrated. (For

This method (EDS/SEM) has also been used for studying photosensitizers in electric tissue

Based on the evidence of aluminum and silicium accumulation in *P. extenta,* we documented the presence of silica minerals in *P. extenta* electric tissue by means of mineralogical techniques (Prado Figueroa *et al*., 2008). It was thought that these compounds could form minerals (i.e., solid inorganic substances with a defined chemical composition and

Fractionation of electric tissue homogenates by differential centrifugation was carried out as described for other tissues (Beaufay and Amar-Costesec, 1976; Amar-Costesec *et al*.,

Fig. 2. Photomicrograph of a cytoplasmic extract of the electric organ from *P. extenta* in

The crossed-polarizer image of a cytoplasmic extract shows SiO2 replacements in grey and

crossed-polarizers, by using a standard polarized light microscope.

white arrangements, with undulatory extinction.

more detail of this method, see Prado Figueroa *et al*., 2008).

(Prado Figueroa & Santiago, 2004).

determined crystallography).

1985)

**2.3 Microcrystalline silica in electric organs** 

using isotonic 3 mM imidazole–HCl-buffered 0.25 M sucrose (pH 7.4). The following fractions were obtained: cytoplasmic extract (E), nuclear fraction (N), large granules (ML), microsomes (P) and supernatant (S). Drops of the fractions were collected on glass slides, dried and mounted in PBS/glycerol. These fractions were inspected using a Leica polarized light microscope (DMLP). This microscope has a polarizer and a switchable analyzer. In mineralogical microscopy, when the light enters an anisotropic mineral, one which transmits light at different rates in different orientations, it is decomposed in two rays, oscillating in two orthogonal planes. This phenomenon is known as birefringence and allows for the identification of each mineral. In this microscope, with a circular graduated stage capable of a 360º rotation, minerals in different positions display their optical properties, such as birefringence colour and extinction position, with crossed polarizers.

All electric organ fractions in crossed-polarizers show SiO2 replacements in grey and white arrangement, with undulatory extinction. The crossed-polarizer image of a cytoplasmic extract shows SiO2 replacements in grey and white arrangements (chalcedony), Fig. 2.

Electric organs without any treatment were also used for X-ray diffraction analysis. Different peaks were obtained from diffractometric analysis; specifically peaks belonged to a quartz (low quartz; Moore and Reynolds, 1997). (See: Prado Figueroa *et al*., 2008).
