The South American gymnotiform knifefish, Brachyhypopomus gauderio is a weakly electric fish species possessing an electric organ (EO). Development of the EO at embryonic and early larval stages has not been extensively studied. B. gauderio is relatively easy to keep in the aquarium and to obtain embryos and therefore could be a good animal model for studying development of the EO. However, little is known about its embryonic development. In this study, we described fertilisation in B. gauderio until early larval stage including 32 stages (180hpf), and determined the larval hatching that was between 3-4 days. The analyses showed a large EO at the ventral side of the muscle (veo). We also discovered another small potential EO-like tissue located at the dorsal side of the muscle (deo) and that has not been previously described. The results also described the derivative electrocytes from the electroprimordium which are separated from the ventral somite muscle in the 3.5dpf during the hatching period. This structure is further separated in the following days. At that time, the first myogenic EO is developed at the border between the ventral somite and the ventral tail fin. During progression of electroprimordium development, the location of the electroprimordium moves down ventrally in the tail fin where newly formed electrocytes appear with multiple nuclei seen in a large syncytial cell. Fluorescent immunohistochemistry showed that MF20 (myosin heavy chain) antibody stained muscles at stages 60hpf, 84hpf, 5.5dpf and 7.5dpf. MF20 also weakly stained the border of electrocytes and the electroprimordium at stages 5.5dpf and 7.5dpf. PAX7 antibody was expressed in the electroprimordium at stages 5.5dpf and 7.5dpf. Moreover, in situ hybridisation staining with scn4aa suggests that cell fate specification of EO may start even earlier at embryonic stage (40hpf). We investigated signalling pathways in EO development using specific inhibitors or activator for key signalling pathways (Bmp, Nodal, Fgf, Retionic Acid and Sonic hedgehog) to the B. gauderio at early stages using in situ hybridisation with scn4aa probe. The results show that Bmp inhibitor, FGF inhibitor and RA suppressed or reduced the scn4aa expression but also reduced the tail and tail fin development. Therefore it was not obvious if these signalling pathways are directly involved in the EO development or if the signals indirectly regulate the EO development by altering cell fates of tail tissues. The SHH inhibitor, cyclopamine treatment at late blastula stage suppressed scn4aa. These results suggest that sonic hedgehog at the late blastula to early gastrula stage has a crucial role in the EO development. Edar morpholino injection suppressed the dorsal EO via deletion of ventral and dorsal fins which was confirmed by the suppression of scn4aa expression in the morphant. This could be a result of the absence of the fin which may possibly be important for activating scn4aa in the dorsal side. To study EO development in the adult fish, the caudal filament was amputated. We observed that tail regeneration occurred during the following two weeks including newly regenerated EO, suggesting that this system would be useful for studying EO development using adult fish, and for learning mechanisms of tissue regeneration in the fish. In the current study, Zn-12 antibody detected a neurogenic EO of electric fish B. gauderio that has not been discovered before. The staining signal with Zn-12 was obvious in the neurogenic EO at adult stage, however the signal at 25dpf was faint and small, and was not detected in the larvae suggesting this organ is specific to juvenile to adult stage. Furthermore, detailed description of the EO structure at early and late stages were conducted using TEM analysis, showing nuclei, ribosomes, glycogen, endoplasmic reticulum, late endosomes and large number of mitochondrion from embryonic electroprimordium to adult EO. At 8.5dpf, the electroprimordium was seen in the front line of the EO moving towards the ventral end of the fin during the EO development. TEM analyses also showed intense myelin layers formed in the neourocytes of neurogenic EO at adult stage (7mpf) considering its important in neourocytes structure. Throughout these studies, we found that the knifefish, B. gauderio embryos and adult fish with the regeneration assay are excellent models for studying development of the electric organ.

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The processing of sensory signals is an important, yet complex task in which a system must extract behaviorally relevant stimulus patterns from a vast array of sensory cues. When a neuron within a major sensory area is presented with a stimulus, one of the important characteristics used to distinguish between types of input is frequency. Often sensory neurons are tuned to narrow stimulus frequency ranges and are thus charged with the processing of subtypes of sensory signals. The weakly electric fish Apteronotus lepthorhynchus senses it's environment through modulations of a self-generated electric field. Two main types of sensory signals can be distinguished based on their frequency patterns. Prey stimuli cause low frequency perturbations of the electric field, while communication signals often result in high frequency signals. Pyramidal neurons in the electrosensory lateral line lobe (ELL) encode the low frequency signals with bursts, while the high frequency signals are relayed with single spikes. This thesis describes how a pyramidal neuron's response patterns can be tuned to specific frequencies by the expression of distinct classes of potassium channels. / I have cloned 3 small conductance (SK) calcium activated potassium channels from cDNA libraries created from the brain of Apteronotus. I have subsequently localized the AptSK channels throughout the brain using both in situ hybridization (AptSK1, 2 & 3) and immunohistochemical (AptSK1 & 2) techniques. The 3 channels showed distinct expression patterns, with the AptSK1 & 2 channels showing a partially overlapping expression pattern, while AptSK3 appears to be expressed in unique areas of the brain. In the ELL AptSK1 & 2 show a partially overlapping expression pattern, appearing in similar pyramidal neurons. However, their distribution within individual cell is unique, with AptSK1 showing a dendritic localization, while AptSK2 is primarily somatic. We have demonstrated that the unique expression pattern of the somatic AptSK2 channel in the ELL coincides with the functional SK currents evaluated through in vitro electrophysiology. Further we have shown that neurons that encode low frequencies do not possess functional SK channels. It thus appears that the presence of the AptSK2 channel subtype can predispose a neuron to respond to specific types of sensory signals. / In an attempt to evaluate if second messengers could modify the AptSK control of frequency tuning I investigated the consequences of muscarinic acetylcholine receptor (mAChR) activation on a pyramidal neurons response patterns. While it had been shown in vivo that mAChR activation increased a pyramidal neuron's response to low frequencies, I have found that this was not due to a decrease in AptSK current, but rather appears to be the result of a down-regulation of an A-type potassium channel. / Taken together the studies that comprise this thesis show how the selective expression of a single potassium channel subtype can control a sensory neurons response to specific environmental cues. The secondary modulation of the A-type current highlights the potential for a second messenger to control a neuron's sensory response through the down-regulation of constitutively expressed potassium current.

Members of voltage-gated potassium channel subfamily 3 (Kv3) have been extensively demonstrated to play a significant role in facilitating function of "fast-firing" neurons in the central nervous system. Kv3.1 and Kv3.3 channels, members of Kv3 channel subfamily, have different distribution profiles on the regional level of brain and on the subcellular level of neurons in mammals and in weakly electric fish, according to mRNA hybridizations in situ and immunohistochemical analysis. In mammals, Kv3.1 channels are expressed in soma, axon and proximal dendrites as well as presynaptic membrane of "fast-firing" neurons. In weakly electric fish (Apteronotus), Kv3.1 channels are distributed in the soma, in the basilar dendrites and in the proximal apical dendrites of pyramidal neurons; on the other hand, Kv3.3 channels are expressed in a larger region: soma, basilar dendrites and entire apical dendrites of these cells. Mechanisms underlying differential subcellular distribution of Kv3.1 and Kv3.3 channels in the apical dendritic compartment of pyramidal neurons are unknown. In order to identify peptide sequences responsible for the differential subcellular localization, I have used Semliki Forest virus as a modified viral expression system (PDE) in vivo to study dendritic targeting mechanisms in the pyramidal neurons of electrosensory lateral line lobe (ELL), where the primary processing for afferent input occurs in the apteronotid electrosensory system.

Partial cDNAs for each of the four known N-methyl-D-aspartate (NMDA) receptor 2 (NMDAR2A-D) subunits have been cloned from the brain of A. leptorhynchus and are found to display a high degree of sequence homology (83--78% amino acid identity) to their mammalian homologues. In situ hybridization experiments reveal that each transcript has a distinct expression pattern in the apteronotid central nervous system (CNS) and is present in a "mosaic" distribution within important cell types of the electrosensory lateral line lobe (ELL). Apt. NMDAR2A transcript is expressed in forebrain regions as well as throughout the pyramidal cell layer (PCL) and granule cell layer (GCL) of the ELL. Apt. NMDAR2B mRNA is enriched in mid- and forebrain structures as well as the PCL and GCL of the ELL. Apt. NMDAR2C transcript is largely restricted to cerebellar regions but is also found in the PCL and GCL of the ELL's medial, centromedial, and centrolateral segments. Apt. NMDAR2D mRNA is expressed in sites of cell proliferation and in a segmental gradient within granule cells of the ELL.

Members of the Dlg/PSD-95 protein family have a modular organization with multiple protein interaction domains and are thought to be important in the organization of synapses. A degenerate primer PCR strategy was used to screen for members of this family in the central nervous system of Apteronotus leptorhynchus, which possesses an electrosensory system that is well-suited for the study of synaptic organization and the subcellular localization of proteins. A. leptorhynchus was found to express at least four Dlg/PSD-95 family genes as in mammals, and the full coding sequences of the homologues of mammalian PSD-95 and SAP102 were determined. In situ hybridization experiments performed for the A. leptorhynchus Dig/PSD-95 family members in brain showed that they have a differential pattern of expression.

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