Early development of the Tsc1 Purkinje cell specific mouse knockouts.

Tsc1 is a gene which expression results in hamartin, a protein involved in regulation of the mTOR1 pathway. Inactivation of Tsc1 gives rise to hyperactivation of the mTOR1 machinery, increased proliferation and growth of cells with subsequent cell degeneration and cell death. In humans, mutations of Tsc1 result in an inherited disorder ­ tuberous sclerosis complex (TSC) with the concomitant multiorgan non­malignant tumors (tubers), epileptic seizures and autistic­like manifestations. General mouse knock­outs, homozygous for the inactivated Tsc1 alleles do not survive and die at early embryonal stages. To circumvent this problem, we utilized the Cre/loxP system and removed Tsc1 specifically in Purkinje cells using the pcp2/L7Cre mouse strain and the Tsc1tmDjk/J strains. Because of the published results showing the autistic­like symptoms after the same crossbred, we have decided to look closer at the early postnatal period of these mutants. Surprisingly no evidence of any behavioral alterations were found, including the ultrasonic vocalizations of newborns. We decided to focus more attention on the interpretation of data, including a more detailed statistical evaluation of our results.

Fear Extinction and Predictive Trait-Like Inter-Individual Differences in Rats Lacking the Serotonin Transporter.

Anxiety disorders are associated with a failure to sufficiently extinguish fear memories. The serotonergic system (5-hydroxytryptamine, 5-HT) with the 5-HT transporter (5-HTT, SERT) is strongly implicated in the regulation of anxiety and fear. In the present study, we examined the effects of SERT deficiency on fear extinction in a differential fear conditioning paradigm in male and female rats. Fear-related behavior displayed during acquisition, extinction, and recovery, was measured through quantification of immobility and alarm 22-kHz ultrasonic vocalizations (USV). Trait-like inter-individual differences in novelty-seeking, anxiety-related behavior, habituation learning, cognitive performance, and pain sensitivity were examined for their predictive value in forecasting fear extinction. Our results show that SERT deficiency strongly affected the emission of 22-kHz USV during differential fear conditioning. During acquisition, extinction, and recovery, SERT deficiency consistently led to a reduction in 22-kHz USV emission. While SERT deficiency did not affect immobility during acquisition, genotype differences started to emerge during extinction, and during recovery rats lacking SERT showed higher levels of immobility than wildtype littermate controls. Recovery was reflected in increased levels of immobility but not 22-kHz USV emission. Prominent sex differences were evident. Among several measures for trait-like inter-individual differences, anxiety-related behavior had the best predictive quality.

Obesity-associated deterioration of the hippocampus is partially restored after weight loss.

OBJECTIVE: Obesity is a multidimensional condition that is treatable by the restoration of a lean phenotype; however, some obesity-related outcomes may persist after weight normalization. Among the organs of the human body, the brain possesses a relatively low regenerative capacity and could retain perturbations established as a result of developmental obesity. Calorie restriction (CR) or a restricted ketogenic diet (KD) are successfully used as weight loss approaches, but their impact on obesity-related effects in the brain have not been previously evaluated. METHODS: We performed a series of experiments in a rat model of developmental obesity induced by a 12-week cafeteria diet, followed by CR to implement weight loss. First, we assessed the impact of obesity on neurogenesis (BrdU incorporation into the hippocampus), cognitive function (water maze), and concomitant changes in hippocampal protein expression (GC/MS-MS, western blot). Next, we repeated these experiments in a rat model of weight loss induced by CR. We also measured mitochondrial enzyme activity in rats after weight loss during the fed or fasting state. This study was extended by additional experiments with restricted KD used as a weight loss approach in order to compare the efficacy of two different nutritional interventions used in the treatment of obesity on hippocampal functions. By using a modified version of the water maze we evaluated cognitive abilities in rats subjected to weight loss by CR or a restricted KD. RESULTS: In this study, obesity affected metabolic processes, upregulated hippocampal NF-κB, and induced proteomic differences which were associated with impaired cognition and neurogenesis. Weight loss improved neurogenesis and enhanced cognition. While the expression pattern of some proteins persisted after weight loss, most of the changes appeared de novo revealing metabolic adjustment by overactivation of citrate synthase and downregulation of ATP synthase. As a consequence of fasting, the activity of these enzymes indicated hippocampal adaptation to negative energy balance during the weight loss phase of CR. Moreover, the effects on cognitive abilities measured after weight loss were negatively correlated with the animal weight measured at the final stage of weight gain. This was alleviated by KD, which improved cognition when used as a weight loss approach. CONCLUSIONS: The study shows that cognition and mitochondrial metabolism in the hippocampus are affected by CR- or KD-induced weight loss.

Preliminary Validation of a Two-Point Estimation Task for the Measurement of Sensory Dissociation in Patients with Chronic Low Back Pain.

OBJECTIVES: Sensory dissociation (SEDI), the discrepancy between perception and actual size or shape of a painful body part, is a frequently observed finding in patients with chronic low back pain. However, the current methods of evaluating SEDI have several limitations, such as a qualitative nature and weak evidence supporting their reliability. In the current study, the reliability of two versions (manual and verbal) of a novel test, a two-point estimation task (TPE), was investigated. METHODS: To perform the manual version of the task, patients estimated the distance between two tactile stimuli delivered to their back using callipers, whereas in the verbal version they verbally reported the estimated distance. RESULTS: The manual version of TPE showed greater interexaminer reliability than the verbal version, and the mean of the two repeated measurements was sufficient for reaching excellent reliability for the pain-free (intraclass correlation coefficient [ICC] = 0.91, 95% confidence interval [CI] = 0.77-0.97) and painful (ICC = 0.86, 95% CI = 0.65-0.94) sides. Intra-examiner reliability was moderate to excellent (ICC = 0.66-0.96) for the manual version performed at the pain-free and painful sides. Distribution, duration, and intensity of pain significantly predicted SEDI and accounted for 42% of the total variance (corrected R2 = 0.42, P < 0.01). CONCLUSIONS: TPE showed higher reliability coefficients compared with tools previously suggested in the literature and can therefore be used clinically and experimentally by one or more examiners. Further research is required to investigate the validity of this new test.

The modification of the ketogenic diet mitigates its stunting effects in rodents.

The high-fat and low-carbohydrate ketogenic diet (HFKD) is extensively studied within the fields of numerous diseases, including cancer and neurological disorders. Since most studies incorporate animal models, ensuring the quality of ketogenic rodent diets is important, both in the context of laboratory animal welfare as well as for the accuracy of the obtained results. In this study we implemented a modification to a commonly used ketogenic rodent chow by replacing non-resorbable cellulose with wheat bran. We assessed the effects of month-long treatment with either the unmodified or the modified HFKD on the growth and development of young male rats. Daily body weight, functional performance, and brain morphometric parameters were assessed to evaluate the influence of both applied diets on rodent development. Our results revealed that the unmodified ketogenic chow induced strong side effects that included weakness, emaciation, and brain undergrowth concomitant to growth inhibition. However, application of the ketogenic chow supplemented with wheat bran suppressed these adverse side effects, which was associated with the restoration of insulin-like growth factor 1 and a decrease in corticosterone levels. We have also shown that the advantageous results of the modified HFKD are not species- or sex-specific. Our data indicate that the proposed HFKD modification even allows for its application in young animals, without causing detrimental side effects.

Genetic Targeting in Cerebellar Purkinje Cells: an Update.

Since the last review paper published in Cerebellum in 2002 [1], there has been a substantial increase in the number of experiments utilizing transgenic manipulations in murine cerebellar Purkinje cells. Most of these approaches were made possible with the use of the Cre/loxP methodology and pcp2/L7 based Cre recombinase expressing transgenic mouse strains. This review aims to summarize all studies which used Purkinje cell specific transgenesis since the first use of mouse strain with Purkinje cell specific Cre expression in 2002.

The point-to-point test: A new diagnostic tool for measuring lumbar tactile acuity? Inter and intra-examiner reliability study of pain-free subjects.

A two-point discrimination test (TPD) is commonly used to investigate lumbar tactile acuity. However, low inter-examiner reliability and difficulties in execution significantly limit its application. Therefore the aim of this study was to compare the inter- and intra-examiner reliability of a new approach, the point-to-point test (PTP), with the TPD. Twenty-one pain-free subjects attended the inter-examiner stage of the study. Eighteen of them were further recruited into an intra-examiner (reproducibility and repeatability) reliability study. PTP was performed on the three points plotted at the L3 spinal level. Point '0' overlapped with the L3 spinous process, from which points '1' and '2' were horizontally separated by 5 and 10 cm, respectively. Participants manually indicated a point previously touched by the examiner, while the distance (error) was measured. Reliability was determined with the intraclass correlation coefficient (ICC2,3). The results revealed good and moderate inter- and intra-examiner reliability at point '1' (ICC2,3 = 0.68-0.84) and good reliability at point '2' (ICC2,3 = 0.84-0.86). At point '0', reliability was moderate to poor (ICC2,3 = 0.13-0.63). TPD was characterised by a poor to moderate level of inter- (ICC2,1 = 0.51; ICC2,3 = 0.56) and intra-examiner reliability (ICC(2,1) = 0.50; ICC2,3 = 0.74). Our findings suggest that PTP is more reliable than TPD at two investigated points at the L3 spinal level. However, further research on PTP validity data is strongly warranted.