Effects Of Melamine On The Excitability Of Rat Hippocampal CA1 Neurons

Melamine, (2,4,6-triamino-s-triazine), is a nitrogen-rich heterocyclic triazine and an industrially synthesized chemical used for a wide variety of applications, and one of those was used primarily in the synthesis of melamine-formaldehyde resins for the manufacture. In our daily life, people are often exposed to products made of melamine-formaldehyde resins such as dishware and kitchenware, which could give off toxic substance when being heated. As the toxic powdered milk scandal protagonist in recent years, there are more widely and deeply researches to investigate its toxicity. It is currently believed that the main target of melamine is urinary system. However, some others studies revealed that melamine also had other biological actions. Such as skin sensibility, liver necrosis, cell apoptosis, cause cancer, change DNA, erythrocyte hemolysis and lead to toxcity in immune and reproductive system.It is well known that infants are more vulnerable to the toxicity of melamine because of underdevelopment of multiple organs, especially kidney and brain. During fetus and infant periods, it plays the most fundamental role in brain development, including increase in neuron number and synaptogenesis which are physiological basis of learning and memory. It has been reported that melamine can pass through the placental barrier and blood-brain barrier. In experimental rats, it was found in different brain regions and could gradually increase with prolongation of expoused time. Our previous study proved that melamine had toxic effect on PC12 cell lines (which was usually used as a neuron model). Thus, our hypothesis was that melamine had certain adverse effect on infants'central nervous system (CNS). It is an important target organ for lots of exogenous compound and now the mechanism of neurotoxic effect was generally regarded as excitability toxicity or oxidative stress as the final commen pathway. Neurons are the basic structural and functional units of the nervous system. They are excitable cells whose excitability is intimately associated with function achievement. Changes of excitability would lead to dysfunction of neurons. Furthermore, various voltage-gated ion channels on neuron membrane are molecular basis of neuron excitability. Many drugs and toxins are target on these ion channels. Therefore, we tested whether melamine had effect on neuron excitability and the underlying ion mechanisms.1. The effect of melamine on spontaneous firing of hippocampal CA1 neurons: melamie increased the frequecy of spontaneous firing in a concentration-dependent mannner, and there were significant differences betwee.control group and melamie group at 5×10-4 and 5×10-5g/mL concentrations. The results suggested that melamine could enhance the excitability of neurons.2. The effect of melamine on evoked action potential of hippocampal CA1 neurons: similarly, melamie increased the frequecy of evoked action potential in a concentration-dependent manner, but it did not change the membrane potential. The results of single action potential revealed that the peak and overshoot value decreased weakly only in 5×10-4g/mL group, while the threshold potential and half-width of action potential were increased in a concentration-dependent manner in 5×10-4, 5×10-5g/ml group.3. The effect of melamine on voltage-gated sodium channels of hippocampal CA1 neurons:melamine could weakly decrease the amplitude of INa in a concentration-dependent manner, and shifted both the steady-state activation and inactivation curves to the hyperpolarizing direction. However, there was no statistical change in the recovery dynamics.4. The effect of melamine on voltage-gated potassium channels of hippocampal CA1 neurons:the results showed that melamine could decrease the amplitude of both IA and IK in a voltage-dependent manner. There were significant changes in IK activation dynamics in a concentration-dependent manner while there were not in IA. What'more, the steady-state inactivation curves of IA were shifted to the left and the slop factors were increased in 5×10-4,5×10-5 g/ml group. There was no significant change in the recovery dynamics.According to the results above, we draw conclusions as follows:melamine could enhance the excitability of neurons in a concentration-dependent manner and the threshold of action potential decreased while the spike half-width prolonged. The effects of melamine on voltage-gated sodium channels could change the activation and inactivation dynamics of INa, which may explain the alternation of threshold of action potential. And the inhibiting effect of melamine on IA and IK may lead to the alternation of frequency and spike half-width which are associated with neuron excitability. Therefore, the effects of melamine on the two main voltage-gated ion channels may be the underlying mechanisms of the enhancing excitability.