Electric catfish hearts are not intrinsically immune to electric shocks.

High voltage electric shocks cause life threatening cardiac injuries such as sudden cardiac standstill or severe myocardial injury. Here, we analysed the physiology of the heart of the strongly electric catfish (Malapterurus beninensis) that stuns prey with high-voltage shocks but is immune to its own, as well as external, high-voltage shocks. Neither a detailed analysis of the electrocardiogram nor the structure of the heart indicated a specialized cardiac conduction system. Using a suitable perfusion system, we discovered that, despite its immunity in vivo, the explanted heart of electric catfish can readily be activated by external electrical currents and is equally sensitive to electric shock-induced arrhythmias as similar-sized goldfish hearts. The surprise thus is that the electric catfish has a vulnerable heart that requires to be protected by highly efficient but presently unknown means.

Connexins evolved after early chordates lost innexin diversity.

Gap junction channels are formed by two unrelated protein families. Non-chordates use the primordial innexins, while chordates use connexins that superseded the gap junction function of innexins. Chordates retained innexin-homologs, but N-glycosylation prevents them from forming gap junctions. It is puzzling why chordates seem to exclusively use the new gap junction protein and why no chordates should exist that use non-glycosylated innexins to form gap junctions. Here, we identified glycosylation sites of 2388 innexins from 174 non-chordate and 276 chordate species. Among all chordates, we found not a single innexin without glycosylation sites. Surprisingly, the glycosylation motif is also widespread among non-chordate innexins indicating that glycosylated innexins are not a novelty of chordates. In addition, we discovered a loss of innexin diversity during early chordate evolution. Most importantly, lancelets, which lack connexins, exclusively possess only one highly conserved innexin with one glycosylation site. A bottleneck effect might thus explain why connexins have become the only protein used to form chordate gap junctions.

Efficient high-voltage protection in the electric catfish.

For thousands of years, starting with detailed accounts from ancient Egypt, the African electric catfish (Malapteruridae) has been renowned for its ability to hunt and to defend itself with powerful electric shocks. Surprisingly, the degree to which electric catfish are protected against their own or external electric shocks, how specific any protection would be to the species-specific waveform and whether a discharging catfish has to actively prepare for the onset of its high-voltage discharges has never been analysed. Here, we used digital high-speed video to record catfish during their own discharges or as they were exposed to external discharges, employing goldfish to carefully calibrate the efficiency of all discharges. Electric catfish show a remarkable degree of protection against high voltages: both self-produced and external electric shocks that heavily affected control goldfish failed to evoke involuntary muscle contraction or to affect sensorimotor processing. Even a commercial electrofishing device, set to efficiently immobilise and narcotise fish, failed to have any effect on the electric catfish. Our findings rule out several protective mechanisms and demonstrate a highly efficient and versatile shielding whose nature is presently unclear.

A Direct Comparison of Different Measures for the Strength of Electrical Synapses.

During the last decades it became increasingly evident that electrical synapses are capable of activity-dependent plasticity. However, measuring the actual strength of electrical transmission remains difficult. Usually changes in coupling strength can only be inferred indirectly from measures such as the coupling coefficient and the coupling conductance. Because these are affected by both junctional and non-junctional conductance, plastic changes can potentially be due to both components. Furthermore, these techniques also require the blocking of chemical transmission, so that processes that involve crosstalk between chemical and electrical synapses will be suppressed. To directly examine the magnitude of errors that can occur, we use dual whole-cell current- and voltage-clamp recordings from the soma of the pair of easily accessible, electrically coupled Retzius cells in the leech to simultaneously determine coupling coefficients, coupling conductances and directly measured gap junctional currents. We present the first direct and comparative analysis of gap junction conductance using all three methods and analyze how each method would characterize the response of gap junctions to serotonin. The traditional coupling coefficients showed severe deficits in assessing the symmetry and strength of electrical synapses. These were reduced when coupling conductances were determined and were absent in the direct method. Additionally, both coupling coefficient and coupling conductance caused large and systematic errors in assessing the size and time course of the serotonin-induced changes of gap junctional currents. Most importantly, both measurements can easily be misinterpreted as implying long-term gap junctional plasticity, although the direct measurements confirm its absence. We thus show directly that coupling coefficients and coupling conductances can severely confound plastic changes in membrane and junctional conductance. Wherever possible, voltage clamp measurements should be chosen to accurately characterize the timing and strength of plasticity of electrical synapses. However, we also demonstrate that coupling coefficients can still yield a qualitatively correct picture when amended by independent measurements of the course of membrane resistance during the experiments.

Recombinant Spider Silk and Collagen-Based Nerve Guidance Conduits Support Neuronal Cell Differentiation and Functionality in Vitro.

Biomaterial scaffolds are under investigation as therapeutic tools to bridge nerve endings following traumatic peripheral nerve injury. The goal is to develop biocompatible nerve guidance conduits (NGCs) with internal guiding structures that promote longitudinally oriented cell migration and regeneration. In the present study, a nonwoven mesh (NWM) made of a recombinant spider silk protein was processed into a tubular structure, ensuring structural integrity of enclosed microfluidics-produced collagen fibers for cell and neurite guidance. The differentiated type of the neuroblastoma X glioma hybrid cell line NG108-15 was used as a model for studying neuronal differentiation on the individual components and on the complete NGC. Differentiated NG108-15 cells grown on recombinant spider silk NWM and collagen fibers formed neuronal networks and synapses. Additionally, whole-cell patch clamp recordings confirmed that all components supported the differentiation of NG108-15 cells into functional neurons. Our NGC demonstrated that tubes made of recombinant spider silk NWM filled with microfluidics-produced collagen fibers are well suited for peripheral nerve repair.

Long-term potentiation in an innexin-based electrical synapse.

Electrical synapses are formed by two unrelated gap junction protein families, the primordial innexins (invertebrates) or the connexins (vertebrates). Although molecularly different, innexin- and connexin-based electrical synapses are strikingly similar in their membrane topology. However, it remains unclear if this similarity extends also to more sophisticated functions such as long-term potentiation which is only known in connexin-based synapses. Here we show that this capacity is not unique to connexin-based synapses. Using a method that allowed us to quantitatively measure gap-junction conductance we provide the first and unequivocal evidence of long-term potentiation in an innexin-based electrical synapse. Our findings suggest that long-term potentiation is a property that has likely existed already in ancestral gap junctions. They therefore could provide a highly potent system to dissect shared molecular mechanisms of electrical synapse plasticity.

Magnetic-activated cell sorting (MACS) can be used as a large-scale method for establishing zebrafish neuronal cell cultures.

Neuronal cell cultures offer a crucial tool to mechanistically analyse regeneration in the nervous system. Despite the increasing importance of zebrafish (Danio rerio) as an in vivo model in neurobiological and biomedical research, in vitro approaches to the nervous system are lagging far behind and no method is currently available for establishing enriched neuronal cell cultures. Here we show that magnetic-activated cell sorting (MACS) can be used for the large-scale generation of neuronal-restricted progenitor (NRP) cultures from embryonic zebrafish. Our findings provide a simple and semi-automated method that is likely to boost the use of neuronal cell cultures as a tool for the mechanistic dissection of key processes in neuronal regeneration and development.