Purkinje cell dysfunction and alteration of long-term synaptic plasticity in fetal alcohol syndrome

Abstract

In cerebellum and other brain regions, neuronal cell death because of ethanol consumption by the mother is thought to be the leading cause of neurological deficits in the offspring. However, little is known about how surviving cells function. We studied cerebellar Purkinje cells in vivo and in vitro to determine whether function of these cells was altered after prenatal ethanol exposure. We observed that Purkinje cells that were prenatally exposed to ethanol presented decreased voltage-gated calcium currents because of a decreased expression of the ␥-isoform of protein kinase C. Longterm depression at the parallel fiber-Purkinje cell synapse in the cerebellum was converted into long-term potentiation. This likely explains the dramatic increase in Purkinje cell firing and the rapid oscillations of local field potential observed in alert fetal alcohol syndrome mice. Our data strongly suggest that reversal of longterm synaptic plasticity and increased firing rates of Purkinje cells in vivo are major contributors to the ataxia and motor learning deficits observed in fetal alcohol syndrome. Our results show that calcium-related neuronal dysfunction is central to the pathogenesis of the neurological manifestations of fetal alcohol syndrome and suggest new methods for treatment of this disorder.calcium ͉ cerebellum ͉ protein kinase ͉ long-term depression ͉ motor learning F etal alcohol syndrome (FAS) is the leading cause of intellectual disability in the Western world with a prevalence of 1 to 1.5 cases per 1,000 live births (1) and a lifetime cost of care of approximately $1.4 million per case. This cost is mainly because of ethanol toxicity in the developing central nervous system, causing intellectual disability, deficits in learning, and fine-motor dysfunction (2).The cerebellum is one of the main targets of in utero ethanol toxicity (3). Within the cerebellum, Purkinje cells (PCs) are highly sensitive to ethanol. PCs constitute the sole output of the cerebellar cortex and thus have a central functional role in integration. All animal models of FAS display a reduction in the number of PCs by Ϸ20% (4), and various authors have proposed that neuronal loss is the only cause of cerebellar deficits in FAS. One study conducted on adult anesthetized rats with FAS led to the conclusion that PCs that survive ethanol administration function normally (5). Thomas et al. (6) found a correlation between total PCs number and motor performance. These experimental data led to the assumption that surviving PCs function normally and that motor coordination impairment in FAS results only from a quantitative defect of PCs. This is crucially important from a therapeutic point of view because very few options exist to replace dead neurons. Many studies have therefore focused on different ways to decrease PC loss in FAS (7,8). However, different models of ataxia that result from PC death per se (pcd mice, SV4 mice, T147 transgenic mice) have demonstrated that considerable neuropathology can occur without the manifestation of a neurologic...

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“…Biocytin was revealed with streptavidin- In vivo extracellular Purkinje cell recordings and eyelid conditioning. In vivo extracellular Purkinje cell recordings and eyelid conditioning in alert mice were performed as previously described (Servais et al, 2007). Briefly, the day before recording sessions, mice were anesthetized with xylido-dihydrothiazin (Rompun; 10 mg/kg; Bayer) and ketamine (Ketalar; 100 mg/ kg; Pfizer).…”

“…Histology was performed as previously described (Servais et al, 2007). Antibodies used were rabbit anti-calbindin-D28k (1/500; Swant), goat polyclonal Kv4.1 antibody (1/50; Santa Cruz Biotechnology), or mouse monoclonal Kv4.2 antibody (10 g/ml) or mouse monoclonal Kv4.3 antibody (100 g/ml) (both gifts from James S. Trimmer, University of California, Davis, Davis, CA).…”

“…Accelerating rotarod (Ugo Basile) was performed as previously described (Servais et al, 2007). Tested groups were weight matched.…”

“…Western blotting was performed as previously described (Servais et al, 2007). Antibodies used were rabbit anti-Kv4.1, Kv4.2, or Kv4.3 (1/200), anti-BDNF (1/500) : sc-546; Santa Cruz Biotechnology], mouse anti-actin (1/ 10,000) (Sigma-Aldrich), mouse anti-polyglutamine 1C2 (1/1000) (Millipore Bioscience Research Reagents), or mouse anti-GAPDH (1/ 10,000) (Sigma-Aldrich).…”

“…Slice preparation, perforated-patch recordings, and study of the parallel fiber-Purkinje cell synapse by electrical stimulation of parallel fibers were performed as previously described (Servais et al, 2007). Slices were perfused with an oxygenated extracellular solution containing the following: 120 mM NaCl, 26 mM NaHCO 3 , 2 mM KCl, 2 mM CaCl 2 , 1.19 mM MgSO 4 , 1.18 mM NaH 2 PO 4 , and 11 mM glucose at room temperature (20 -25°C).…”