Effects Of CeCl₃ On Voltage-gated Calcium Channels In Rat Hippocampal Pyramidal Neurons

Objective:Cerium?Ce?is the most abundant light rare earth element in nature with lively chemistry.The polyanions Ce³⁺ could react with almost all the elements and organizational composition,and then enriched in the body.With the application of cerium and its compounds,the toxic effects of Ce³⁺ in multiple organs and system have been extensively studied[1-3].The studies showed Ce³⁺ has neurotoxicity[3-6] which could damage learning,memory and cognitive function in infants and young children,but the mechanism of Ce³⁺ neurotoxicity is still not clear.Because of the similar ion radius with Calcium(Ca²⁺),Ce³⁺ may substitute Ca²⁺ binding to biological macromolecules and interfere the normal function of cells and the intracellular calcium homeostasis.Ca²⁺ is closely related to learning and memory function.Appropriate Ca²⁺ concentration is the basis of neuronal growth,development and synaptic plasticity[11],and the voltage-gated calcium channels plays an important role in regulating Ca²⁺ concentration in neurons[12-14].Therefore,we try to study the effect of Ce³⁺ on voltage-gated calcium channels and calcium oscillation in hippocampal pyramidal neurons by patch clamp technique,in order to providing the theoretical basis and basis for cerium hippocampal neurotoxicity mechanism.Methods:The acute isolated brain slices were prepared from 13-15 day SD rats.The slices were treated with CeCl₃.Whole-cell patch-clamp technique on voltage clamp mode was used to collect Voltage-gated calcium channels current and acetylcholine?Ach?-induced calcium-activated chloride current in the CA1 pyramidal neurons before and after exposure Ce³⁺.The effect of Ce³⁺ on the voltage-gated calcium channel was evaluated by analyzing the change of calcium channel current,I-V curve,steady-state inactivation curve and steady-state activation curve before and after exposure.The changes of calcium-activated chloride current were analyzed to reflect the effect of cerium on Ach induced calcium oscillations in hippocampal pyramidal neurons.Results: 1.The characteristics of high-voltage and low-voltage activated calcium channel currents in pyramidal neurons of hippocampal CA1 regionThe current density of high-voltage activated Ca²⁺ channels in the hippocampal CA1 pyramidal neurons was-15.45±1.94 pA/pF,and 0.2 mmol/L CdCl₂ could completely block IHAV?n = 6?.The low-voltage activated calcium channel current density was-4.84±0.55 p A/pF,and 0.2 mmol/L NiCl₂ inhibit 63.3±7.4% of ILVA?n = 6?.2.The effects of cerium chloride on voltage-gated calcium channel currents in pyramidal neuronsAfter the formation of whole cell model,the slices was perfused with normal artificial cerebrospinal fluid for 5min,and then perfused with artificial cerebrospinal fluid containing 0,5,10,50,100 ?mol/L cerium chloride for 15 min.It was observed that cerium chloride could inhibit IHVA on the pyramidal neurons?P <0.05,n = 8?,but having no effect on ILVA of pyramidal neurons?P>0.05,n = 8?.L-type calcium channel blocker nifedipine can reduce the effect of cerium chloride on IHVA.3.Effect of cerium chloride on the characteristics of high-voltage activated calcium channelThe maximum current density of high-voltage calcium decreased from-15.45± 1.94 pA/pF to-8.74±1.58 pA/pF?P <0.05,n = 8?.The density I-V curve has no significant effect on the shape after exposure.The steady-state activation curves and inactivation curves were fitted and there was no significant difference in the halfactivation voltage,half-inactivation voltage and slope rate before and after exposure to cerium chloride?P> 0.05,n = 10?.4.Effect of cerium chloride on Ach-induced ccalcium oscillation10 nmol/L Ach could induce calcium oscillations on pyramidal neurons in CA1 region.50 ?mol/L cerium chloride significantly reduced Ach-induced pyramidal neuronal calcium oscillations,with a normalized net charge of 1.02±0.02?P <0.05,n = 6?.Exposuring to the high-voltage activated calcium channel blocker CdCl2?0.2 mmol/L?,it was observed that the calcium oscillations were significantly decreased,the normalized net charge reduced to 0.30±0.05?n = 6?,after combining with 50?mol/L cerium chloride,the normalized net charge reduced to 0.33±0.04?n = 6?,there was no significantly difference between two groups.Conclusion: 1.Ce³⁺ exposure suppressed high-voltage gated calcium current,but has no significant effect on low-voltage gated calcium current.2.Ce³⁺ exposure reduced the activity of calcium channels in pyramidal neurons,but has no significant effect on the ion channel gating characteristics.3.Ce³⁺ could act on high-voltage activated calcium channels to inhibit Ach-induced calcium on pyramidal neurons in hippocampal.