Insulin and the insulin-like growth factors (IGFs) may directly affect the development of the nervous system. NGF, IGF-II, and insulin's effects on neurite formation and neuronal survival were studied in peripheral ganglion cell cultures from chick embryos. Neurite outgrowth was enhanced in a dose-dependent manner by insulin and IGF-II in sympathetic cell cultures. The half-maximally effective concentration, ED50, was about 0.4-0.6 nM for both polypeptides, and concentrations as low as 10 pM were active. However, in sensory neurons the ED50 for neurite outgrowth was about 30 nM for insulin and 0.1 nM for IGF-II, suggesting that these factors may have selective effects in different neuronal tissues. Neither serum nor the presence of non-neuronal cells was required for the response in sympathetic neurons. The specific anti-NGF antiserum inhibited the neurite outgrowth response to NGF but not to insulin nor IGF-II. Insulin and IGF-II additionally supported survival of sensory and sympathetic neurons; however, insulin was not as efficacious as NGF. The combination of high concentrations of NGF and insulin was no better than NGF alone in supporting sympathetic cell survival, or neurite outgrowth. This indicates that insulin acts on the same, or a subpopulation, of NGF-responsive neurons. These results support the hypothesis that insulin and its homologs belong to a broad family of neuritogenic polypeptides.

LAs can cause rapid cell death, which is primarily due to necrosis. Lidocaine and bupivacaine can trigger apoptosis with either increased time of exposure or increased concentration. These effects might be related to postoperative neurologic injury. Lidocaine, linked to the highest incidence of transient neurological symptoms, was not the most toxic LA, whereas bupivacaine, a drug causing a very low incidence of transient neurological symptoms, was the most toxic LA in our cell model. This suggests that cytotoxicity-induced nerve injury might have different mechanisms for different LAs and different target(s) other than neurons.

The effect of glycosylation on Kv1.1 potassium channel function was investigated in mammalian cells stably transfected with Kv1.1 or Kv1.1N207Q. Macroscopic current analysis showed that both channels were expressed but Kv1.1N207Q, which was not glycosylated, displayed functional differences compared with wild‐type, including slowed activation kinetics, a positively shifted V1/2, a shallower slope for the conductance versus voltage relationship, slowed C‐type inactivation kinetics, and a reduced extent of and recovery from C‐type inactivation. Kv1.1N207Q activation properties were also less sensitive to divalent cations compared with those of Kv1.1. These effects were largely due to the lack of trans‐Golgi added sugars, such as galactose and sialic acid, to the N207 carbohydrate tree. No apparent change in ionic current deactivation kinetics was detected in Kv1.1N207Q compared with wild‐type. Our data, coupled with modelling, suggested that removal of the N207 carbohydrate tree had two major effects. The first effect slowed the concerted channel transition from the last closed state to the open state without changing the voltage dependence of its kinetics. This effect contributed to the G‐V curve depolarization shift and together with the lower sensitivity to divalent cations suggested that the carbohydrate tree and its negatively charged sialic acids affected the negative surface charge density on the channel's extracellular face that was sensed by the activation gating machinery. The second effect reduced a cooperativity factor that slowed the transition from the open state to the closed state without changing its voltage dependence. This effect accounted for the shallower G‐V slope, and contributed to the depolarized G‐V shift, and together with the inactivation changes it suggested that the carbohydrate tree also affected channel conformations. Thus N‐glycosylation, and particularly terminal sialylation, affected Kv1.1 gating properties both by altering the surface potential sensed by the channel's activation gating machinery and by modifying conformational changes regulating cooperative subunit interactions during activation and inactivation. Differences in glycosylation pattern among closely related channels may contribute to their functional differences and affect their physiological roles.

No abstract

Highly purified sodium channel protein from the electric eel, Electrophorus electricus, was reconstituted into liposomes and incorporated into planar bilayers made from neutral phospholipids dissolved in decane. The purest sodium channel preparations consisted of only the large, 260-kD tetrodotoxin (TTX)-binding polypeptide. For all preparations, batrachotoxin (BTX) induced long-lived single-channel currents (25 pS at 500 mM NaCl) that showed voltage-dependent activation and were blocked by TTX. This block was also voltage dependent, with negative potentials increasing block. The permeability ratios were 4.7 for Na+:K + and 1.6 for Na+:Li +. The midpoint for steady state activation occurred around -70 mV and did not shift significantly when the NaCI concentration was increased from 50 to 1,000 mM. Veratridine-induced single-channel currents were about half the size of those activated by BTX. Unpurified, nonsolubilized sodium channels from E. electricus membrane fragments were also incorporated into planar bilayers. There were no detectable differences in the characteristics of unpurified and purified sodium channels, although membrane stability was considerably higher when purified material was used. Thus, in the eel, the large, 260-kD polypeptide alone is sufficient to demonstrate single-channel activity like that observed for mammalian sodium channel preparations in which smaller subunits have been found.