The results of these tests are of interest because activity-induced slowing of spike propagation can reduce the maximum spike frequency generated by axons (Partida et al., 2018) and this would modulate synaptic transmission to neurons in sub-cortical brain regions (Singer et al., 1972; Usrey et al., 1998). Materials and methods Animals Adult Long-Evans rats (female; P60-P120; 150-250g) were obtained from a commercial supplier (Envigo Bioproducts) and housed in standard cages at 23C on a 12-h/12-h light/dark cycle. subtype of K+ channel that associates with optic nerve CaMKII has not been identified. The present study begins to address these questions in four steps. First, we Toceranib (PHA 291639, SU 11654) ask if 4AP slows spike repolarization, as would be expected if 4AP reduces outward K+ current as in other tissues (Storm, 1987). Second, we ask if 4AP slows spike propagation, as proposed for KN-93 (Partida et al., 2018). Third, we ask if antibodies directed against 4AP-sensitive and CaMKII-regulated K+ channels bind to optic nerve axons. Fourth, we ask if CaMKII co-immunoprecipitates with the subtype of K+ channel we immunolocalize in optic nerve axons. Answering the third and fourth questions depends on answers we obtain to the first and second questions. However, answering the first two questions by use of compound action potentials is hindered by two properties Rabbit polyclonal to AKT1 of their waveforms. One is that optic nerve compound action potentials display multiple peaks (cf., Bishop et al., 1953). The second is that these peaks partially overlap (Stys et al., 1991). Because each peak is formed by spikes propagating in multiple axons at similar but not identical speeds, the rate at which each peak rises and falls is weighted by increases and decreases in the number of spikes propagating at different speeds. Because the cardinal peak starts from baseline, and the next peak starts to rise before the cardinal peak returns to baseline, the most rapidly propagating spikes can be timed by the base of the cardinal peak, but the start of each post-cardinal peak is hidden. We therefore record spikes in the present study with extracellular electrodes that can resolve their rate of depolarization separately from their repolarization, and that can more clearly show the time that elapses as spikes propagate between stimulating and recording electrodes. The results of these tests are of interest because activity-induced slowing of spike propagation can reduce the maximum spike frequency generated by axons (Partida et al., 2018) and this would modulate synaptic transmission to neurons in sub-cortical brain regions (Singer et al., 1972; Usrey et al., 1998). Materials and methods Animals Adult Long-Evans rats (female; P60-P120; 150-250g) were obtained from a commercial supplier (Envigo Bioproducts) and housed in standard cages at 23C on a 12-h/12-h light/dark cycle. For the experiments reported here, each rat was anesthetized by intraperitoneal ketamine and xylazine (70C100 mg/kg and 5C10 mg/kg, respectively; see below for the source of chemicals used in this study) and decapitated. The optic nerves were dissected and electrophysiologically recorded from, immunohistochemically stained, and western blotted, as described below. The results presented here are based on measurements obtained from a total of 35 rats (3 for spike recordings, 2 for immunohistochemistry, and 30 for western blots). All animal care and experimental protocols were approved by the Animal Use and Care Administrative Advisory Committee of the University of California, Davis. Spike recordings and analysis For electrophysiological recordings, we isolated the retina from the Toceranib (PHA 291639, SU 11654) eye without removing the Toceranib (PHA 291639, SU 11654) optic nerve from the retina, laid the retina ganglion cell-side down on a multi-electrode array (MEA; Meister et al., 1994), and held the cut end of the optic nerve in a glass suction electrode. This allowed us to stimulate the optic nerve, record action potentials intraretinally (Marchiafava, 1976), and recognize the triphasic waveform of axonal spikes (Figures.