Rat sciatic nerve schwann cell microcultures: Responses to mitogens and production of trophic and neurite-promoting factors

During embryonic development and in response to injury, the growing axons of peripheral neurons may influence the migration and proliferation of Schwann cells which, in return, may present neurons with a critical supply of factors required for neuronal survival, growth and differentiation. The identification and characterization of agents influencing the proliferation of Schwann cells as well as Schwann cell production of factors affecting neurons is greatly facilitated by the use of in vitro techniques. We describe here a simplified method of obtaining large numbers of purified neonatal rat sciatic nerve Schwann cells for use in generating large numbers of replicate microcultures. We then illustrate the use of these microcultures to examine Schwann cell: i) morphology and survival; ii) proliferation; and iii) production of neuronotrophic and neurite-promoting activities. We report that rat Schwann cells in microculture proliferate in response to serum, laminin and fibronectin, cholera toxin, and chick embryo parasympathetic ciliary neurons. Also, extracts of Schwann cell microcultures contain independently regulated activities which support the survival and neurite outgrowth of peripheral ganglionic neurons.Special issue dedicated to Dr. Paola S. Timiras     

Describing the ideal Australian companion dog

Dogs have had a long association with humans and are believed to be the first domesticated animal species. Many breeds of dog exist today which vary considerably in physical appearance and temperament. These differences have arisen primarily from selective pressures imposed by humans to create dogs suitable for various working roles. Nowadays, however, few breeds undertake the work they were once bred for; rather dogs are kept primarily as companions. With differing lifestyles and an increase in urban living it is necessary to identify what constitutes an ideal dog in the present day. This study identifies the characteristics important to the Australian public in their “ideal dog”. To determine this, 877 participants (79.8% female) aged 18–82 years (mean = 34.3, SD = 14.5) were surveyed. A number of behavioural and physical characteristics were identified as important to Australians. These included dogs being medium sized, short haired, de-sexed, safe with children, fully housetrained, friendly, obedient and healthy. Participants also wanted their ideal dog to come when called, not to escape from their property, to enjoy being petted and to display affection to their owners. Desirable behavioural characteristics were grouped using Principal Component Analysis into five factors, labelled calm/compliant, sociable/healthy, energetic/faithful/protective, socially acceptable, and non-aggressive. Together these accounted for 45.7% of the total variance. Independent-samples t-tests revealed significant differences in importance of the components for men versus women, dog owners versus non-owners and whether participants lived with children or not. Women preferred a dog who is calm/compliant [t(870) = −2.33, P = 0.02], sociable/healthy [t(870) = −2.57, P = 0.01] and non-aggressive [t(870) = −2.67, P = 0.008] while men preferred a dog which is energetic/faithful/protective [t(870) = 3.09, P = 0.002]. Overall, however there were also many commonalities. Breeding animals able to tolerate the stresses and demands of today's requirements, training them to behave appropriately, and educating pet dog owners about the characteristics of different dogs and the need for realistic expectations about dog behaviour is likely to help reduce the incidence of problem behaviours, such as separation anxiety, destructiveness and aggression. It is also likely to increase owner satisfaction and reduce the number of dogs relinquished to shelters.     

The denatured state of N-PGK is compact and predominantly disordered

The organisation of the structure present in the chemically denatured N-terminal domain of phosphoglycerate kinase (N-PGK) has been determined by paramagnetic relaxation enhancements (PREs) to define the conformational landscape accessible to the domain. Below 2.0 M guanidine hydrochloride (GuHCl), a species of N-PGK (denoted I_b) is detected, distinct from those previously characterised by kinetic experiments [folded (F), kinetic intermediate (I_k) and denatured (D)]. The transition to I_b is never completed at equilibrium, because F predominates below 1.0 M GuHCl. Therefore, the ability of PREs to report on transient or low population species has been exploited to characterise I_b. Five single cysteine variants of N-PGK were labelled with the nitroxide electron spin-label MTSL [(1-oxyl-2,2,5,5-tetramethyl-3-pyrroline-3-methyl)methanesulfonate] and the denaturant dependences of the relaxation properties of the amide NMR signals between 1.2 and 3.6 M GuHCl were determined. Significant PREs for I_b were obtained, but these were distributed almost uniformly throughout the sequence. Furthermore, the PREs indicate that no specific short tertiary contacts persist. The data indicate a collapsed state with no coherent three-dimensional structure, but with a restricted radius beyond which the protein chain rarely reaches. The NMR characteristics of I_b indicate that it forms from the fully denatured state within 100 μs, and therefore a rapid collapse is the initial stage of folding of N-PGK from its chemically denatured state. By extrapolation, I_b is the predominant form of the denatured state under native conditions, and the non-specifically collapsed structure implies that many non-native contacts and chain reversals form early in protein folding and must be broken prior to attaining the native state topology.     

The 5-Choice Continuous Performance Test: Evidence for a Translational Test of Vigilance for Mice

Background

Attentional dysfunction is related to functional disability in patients with neuropsychiatric disorders such as schizophrenia, bipolar disorder, and Alzheimer''s disease. Indeed, sustained attention/vigilance is among the leading targets for new medications designed to improve cognition in schizophrenia. Although vigilance is assessed frequently using the continuous performance test (CPT) in humans, few tests specifically assess vigilance in rodents.

Methods

We describe the 5-choice CPT (5C-CPT), an elaboration of the 5-choice serial reaction (5CSR) task that includes non-signal trials, thus mimicking task parameters of human CPTs that use signal and non-signal events to assess vigilance. The performances of C57BL/6J and DBA/2J mice were assessed in the 5C-CPT to determine whether this task could differentiate between strains. C57BL/6J mice were also trained in the 5CSR task and a simple reaction-time (RT) task involving only one choice (1CRT task). We hypothesized that: 1) C57BL/6J performance would be superior to DBA/2J mice in the 5C-CPT as measured by the sensitivity index measure from signal detection theory; 2) a vigilance decrement would be observed in both strains; and 3) RTs would increase across tasks with increased attentional load (1CRT task<5CSR task<5C-CPT).

Conclusions

C57BL/6J mice exhibited superior SI levels compared to DBA/2J mice, but with no difference in accuracy. A vigilance decrement was observed in both strains, which was more pronounced in DBA/2J mice and unaffected by response bias. Finally, we observed increased RTs with increased attentional load, such that 1CRT task<5CSR task<5C-CPT, consistent with human performance in simple RT, choice RT, and CPT tasks. Thus we have demonstrated construct validity for the 5C-CPT as a measure of vigilance that is analogous to human CPT studies.