Ca²⁺ homeostasis is crucial for the survival of most if not all multicellular organisms. In fresh water teleost species such as the zebrafish (Danio rerio), the hydroxyapatite (Ca₁₀(PO₄)₆(OH)₂)-rich elasmoid scales that comprise the dermal skeleton, represent a significant internal reservoir of Ca²⁺. However, little is known about how scale tissue contributes to Ca²⁺ homeostasis in fish. Using an ultra-sensitive scanning ion-selective technique (SIET) in conjunction with Ca²⁺-specific ion-selective microelectrodes, I have demonstrated that in vitro, scales can generate short-term (i.e., minutes-to-hours) Ca²⁺ fluxes in response to different [Ca²⁺] in the external medium. When challenged with hypocalcemic ([Ca²⁺]_ext. = 0.01 mM) and hypercalcemic ([Ca²⁺]_ext. = 3.0 mM) media, they ge...[ Read more ]

Ca²⁺ homeostasis is crucial for the survival of most if not all multicellular organisms. In fresh water teleost species such as the zebrafish (Danio rerio), the hydroxyapatite (Ca₁₀(PO₄)₆(OH)₂)-rich elasmoid scales that comprise the dermal skeleton, represent a significant internal reservoir of Ca²⁺. However, little is known about how scale tissue contributes to Ca²⁺ homeostasis in fish. Using an ultra-sensitive scanning ion-selective technique (SIET) in conjunction with Ca²⁺-specific ion-selective microelectrodes, I have demonstrated that in vitro, scales can generate short-term (i.e., minutes-to-hours) Ca²⁺ fluxes in response to different [Ca²⁺] in the external medium. When challenged with hypocalcemic ([Ca²⁺]_ext. = 0.01 mM) and hypercalcemic ([Ca²⁺]_ext. = 3.0 mM) media, they generate outward and inward Ca²⁺ fluxes, respectively; whereas exposure to isocalcemic ([Ca²⁺]_ext. = 1.5 mM Ca²⁺) medium resulted in little-to-no flux generation. When challenged by different calcemic conditions, larger fluxes were consistently recorded on the episquamal (outer) side of the scale, while the hyposquamal (inner) side appears to be less dynamic with respect to Ca²⁺ fluxes. These responses occurred in the absence of any exogenous calciotropic hormones, thus suggesting that the scales cells of zebrafish possess an extracellular Ca²⁺ sensor. The Ca²⁺ efflux generated by scales in hypocalcemic conditions appears to be cell-dependent as it was abolished by treatment with potassium cyanide treatment or by dehydration. Scales were immunolabelled with antibodies to Zns5 and cathepsin K (CtsK), in an attempt to identify the osteoblasts (i.e., cells responsible for Ca²⁺ mineralization in scales) and osteoclasts (i.e., cells responsible for Ca²⁺ mobilization in scales), respectively. Both antibodies seemed to label the same populations of cells on both the episquamal and hyposquamal side of the scales. For example, both antibodies labelled elongated cells residing in the circuli and in the radial grooves on the episquamal side of scales. In addition, both antibodies labelled the uniform squamous layer of cells covering the entire hyposquamal side of the scale, although Zns5 labelling of these cells was most prominent in the plasma membrane, whereas Ctsk labelled distinct regions of the cytosol. To investigate whether these osteoblasts and/or osteoclasts are able to respond to well-known calciotropic hormones, a RT-PCR study of the mRNA transcripts from zebrafish scale tissues was carried out. The results indicate that receptors for parathyroid hormone (Pth), calcitonin (CT), and estrogen are present in scale tissues. Furthermore, treatment of the scales with CT in medium containing physiological levels of [Ca²⁺] (i.e., 1.5 mM Ca²⁺) showed a Ca²⁺ influx into the scales that increased in a dose-dependent manner. On the other hand, treatment with Pth or 17β-estradiol did not have a significant effect on altering the normal Ca²⁺ flux generated by scales bathed in physiological measuring medium. The short-term, relatively fast Ca²⁺ flux responses generated by scale tissues reacting to different external calcemic conditions and/or hormonal signals, resembles the short-term homeostatic response reported in mammalian bone. In the case of mammalian bone, the short-term response occurs in a faster time-frame (minutes-to-hours) than that reported for the long-term “bone-remodelling” response (hours-to-days) that involves bone Ca²⁺ mobilization or mineralization. My new data thus suggest that zebrafish scales also possess a short-term response mechanism to help regulate plasma [Ca²⁺] following specific calcemic challenges. This adds to a growing body of evidence that supports the suggestion that fish scales might be suitable for use as an inexpensive non-mammalian model to study bone development, disease, damage, repair, and regeneration.