The science of research: The principles underlying the discovery of cognitive and other biological mechanisms

Studies of cognitive function include a wide spectrum of disciplines, with very diverse theoretical and practical frameworks. For example, in Behavioral Neuroscience cognitive mechanisms are mostly inferred from loss of function (lesion) experiments while in Cognitive Neuroscience these mechanisms are commonly deduced from brain activation patterns. Although neuroscientists acknowledge the limitations of deriving conclusions using a limited scope of approaches, there are no systematically studied, objective and explicit criteria for what is required to test a given hypothesis of cognitive function. This problem plagues every discipline in science: scientific research lacks objective, systematic studies that validate the principles underlying even its most elemental practices. For example, scientists decide what experiments are best suited to test key ideas in their field, which hypotheses have sufficient supporting evidence and which require further investigation, which studies are important and which are not, based on intuitions derived from experience, implicit principles learned from mentors and colleagues, traditions in their fields, etc. Philosophers have made numerous attempts to articulate and frame the principles that guide research and innovation, but these speculative ideas have remained untested and have had a minimal impact on the work of scientists. Here, I propose the development of methods for systematically and objectively studying and improving the modus operandi of research and development. This effort (the science of scientific research or S2) will benefit all aspects of science, from education of young scientists to research, publishing and funding, since it will provide explicit and systematically tested frameworks for practices in science. To illustrate its goals, I will introduce a hypothesis (the Convergent Four) derived from experimental practices common in molecular and cellular biology. This S2 hypothesis proposes that there are at least four fundamentally distinct strategies that scientists can use to test the connection between two phenomena of interest (A and B), and that to establish a compelling connection between A and B it is crucial to develop independently confirmed lines of convergent evidence in each of these four categories. The four categories include negative alteration (decrease probability of A or p(A) and determine p(B)), positive alteration (increase p(A) and determine p(B)), non-intervention (examine whether A precedes B) and integration (develop ideas about how to get from A to B and integrate those ideas with other available information about A and B). I will discuss both strategies to test this hypothesis and its implications for studies of cognitive function.     

Evidence that androgenic and estrogenic metabolites contribute to the effects of dehydroepiandrosterone on cognition in postmenopausal women

Prior studies of the effects of dehydroepiandrosterone (DHEA) on cognition have produced complex and inconsistent results. We hypothesize that these results may arise, in part, because of DHEA's metabolism into estrogens and androgens that produce opposing effects on cognition. Our study administered 50 mg of oral DHEA daily for 4 weeks in a placebo-controlled crossover design to six postmenopausal women. We measured blood levels of androgens (total testosterone, free testosterone, DHEA, DHEAS), estrogens (estradiol, estrone), and cognitive performance on recognition memory, perceptual identification, digit span memory, and visual attentional vigilance under both drug and placebo conditions. Multiple regression models incorporating the factors of age and body mass index (BMI) were used to ascertain the relation between sex steroids and cognitive performance. Our results demonstrated that estrogens produced a positive effect on recognition memory, while androgens produced a negative effect. This pattern reversed in perceptual identification with estrogens producing a negative effect and androgens producing a positive effect. In addition, BMI produced a negative effect on digit span memory, age produced a negative effect on perceptual identification, and androgens produced a negative effect on visual attentional vigilance. These results help, in part, to explain DHEA's complex effects on cognition. The diverse effects of sex steroids across tasks underscore the importance of identifying the specific cognitive mechanisms influenced by sex steroids and emphasizes that one should not expect sex steroids to produce homogeneous effects across cognitive tasks.     

The learning ability and memory retention of broiler breeders: 1 effects of reduced balanced protein diet on reward based learning

Learning and mental abilities of farm animals are important for their adaptation to new environments and could serve as an indicator for welfare and performance. This study assessed the learning ability and memory retention of broiler breeders through a T-maze test with a reward v. no reward set-up. Feed supplemented with or without meal worms served as a reward. Two trials were conducted with either parent stock (PS) breeders or pure line A (PL) breeders and with the same dietary treatment namely a control (C) group fed with standard commercial diets and a reduced balanced protein (RP) group fed with RP diets (25% reduction of CP and amino acids). To maintain similar target BW, the RP group received on average 10% more feed. A tonic immobility (TI) test was performed to estimate the fearfulness of PS breeders. Most breeders were capable of completing the T-maze tests, but the C group needed less time compared with the RP group when their own feed was given as a reward. However, when meal worms were provided as an extra incentive, the RP group completed the maze significantly faster than the C group. Compared to the C breeders, the RP breeders remained longer in a TI state, indicating a higher level of fearfulness. Long-term memory retention in the T-maze test was observed in both groups. It is concluded that the RP diet had no influences on the learning ability of the reward v. no reward discrimination test and its memory retention of broiler breeders. The increased amount of time the RP breeders needed to solve the test was probably due to a higher sense of fear. Furthermore, the RP diet enhanced the motivation of breeders to obtain an alternative feed such as meal worms.     

The Sensitivity of Cognitive Processes to the Inhomogeneity of Natural Magnetic Fields

Three experiments were carried out to study the influence of inhomogeneity of natural magnetic field (MF) on animal cognition. Wistar rats (n = 90) were placed in a complicated problem environment, in which they were to form food-operant behavior under conditions of natural MF (Cond. 1) and MFs produced by iron objects (Cond. 2) or by magnets (Cond. 3). Unlike the control group (Cond. 1), all rats in Cond. 2 and 3 were unable to form operant behavior. Weak MF caused both locomotor and emotional depression, and there was no exploratory activity shown during 6 sessions. Brief external stimulation removed locomotor depression, and animals formed operant behavior similar to latent learning (Cond. 2) or “insight” (Cond. 3). Performance efficiency was lower while the level of stress manifestation was higher in Cond. 2 and 3 than in the control at the stage of stabilization. It has been proposed that MF reduces the activity of brain motivation centers and prevents the development of complicated forms of cognitive activity.     

Haploinsufficiency of the arginine-vasopressin gene is associated with poor spatial working memory performance in rats

Behavioral pharmacological studies have implicated a role for the neurophysin arginine-vasopressin in learning and memory. Vasopressin, and its analogues, can produce either improvements or impairments in mnemonic functions, effects that depend upon the agent administered, the memory process measured and the task employed. As recent data have implicated vasopressin in regulating the cognitive functions of the prefrontal cortex, we sought to determine whether changes in vasopressinergic tone would affect a form of memory that is dependent upon this brain region. To that end, we used a genetic approach to examine how haploinsufficiency of the vasopressin gene affects working memory performance. Specifically, we tested a naturally occurring null-mutant rat on an operant delayed-non-match-to-position task. Male and female heterozygous and wild-type rats were trained to perform this working memory task, and the effects of varying the delay across which they had to maintain task information were systematically varied. Although vasopressin-deficient rats omitted fewer trials and completed trials more quickly, they exhibited delay-dependent deficits of choice accuracy. The genotype effects were not modified by sex. Collectively, these data indicate that even partial vasopressin deficiency can trigger deficits of spatial working memory performance and add to the growing body of results supporting a regulatory control of neocortical-dependent cognitive functions by this neurohormone.     

The role of awareness and working memory in human transitive inference

The human ability to perform transitive inference (TI) is an area of debate from a neurocognitive standpoint. Some studies emphasize a stimulus driven medial-temporal lobe process [Preston, A.R., Shrager, Y., Dudukovic, N.M., Gabrieli, J.D., 2004. Hippocampal contribution to the novel use of relational information in declarative memory. Hippocampus 14, 148-152; Titone, D., Ditman, T., Holzman, P., Eichenbaum, H., Levy, D., 2004. A transitive inference test of relational memory in schizophrenia. Schizophr. Res. 68, 235-247; Van Elzakker, M., O'Reilley, R., Rudy, J., 2003. Transivity, flexibility, conjenctive representation and the hippocampus: an empirical analysis. Hippocampus 13, 334-340] while others emphasize a higher-level frontal lobe strategy that requires the flexible maintenance of information in working memory [Waltz, J., Knowlton, B., Holyoak, K., Boone, K., Mishkin, F., de Menedezes Santos, M., Thomas, C., Miller, B., 1999. A system for relational reasoning in human prefrontal cortex. Psychol. Sci. 10, 119-125]. In two experiments we investigated when and how adults employ different cognitive strategies during TI by evaluating the interaction between task instructions and individual differences in working memory capacity. Participants engaged in a paired discrimination task involving a 6-unit TI hierarchy and were either prior aware, prior unaware or serendipitously aware of the hierarchical relationship among stimulus items. Both prior aware participants and serendipitously aware participants were more likely to engage in a logic-based strategy compared to unaware participants who relied upon stimulus-driven strategies. Individual differences in working memory were associated with the acquisition of awareness in the serendipitously aware group and with the maintenance of awareness in the prior aware group. These findings suggest that the capacity for TI may be supported by multiple neurocognitive strategies, and that the specific strategy employed is dependent upon both task- and participant-related factors.     

Androgens with activity at estrogen receptor beta have anxiolytic and cognitive-enhancing effects in male rats and mice

Testosterone (T) and its metabolites may underlie some beneficial effects for anxiety and cognition, but the mechanisms for these effects are unclear. T is reduced to dihydrotestosterone (DHT), which can be converted to 5α-androstane,3α,17β-diol (3α-diol) and/or 5α-androstane-3β,17β-diol (3β-diol). Additionally, T can be converted to androstenedione, and then to androsterone. These metabolites bind with varying affinity to androgen receptors (ARs; T and DHT), estrogen receptors (ERβ; 3α-diol, 3β-diol), or GABA_A/benzodiazepine receptors (GBRs; 3α-diol, androsterone). Three experiments were performed to investigate the hypothesis that reduced anxiety-like and enhanced cognitive performance may be due in part to actions of T metabolites at ERβ. Experiment 1: Gonadectomized (GDX) wildtype and ERβ knockout mice (βERKO) were subcutaneously (SC) administered 3α-diol, 3β-diol, androsterone, or oil vehicle at weekly intervals, and tested in anxiety tasks (open field, elevated plus maze, light–dark transition) or for cognitive performance in the object recognition task. Experiment 2: GDX rats were administered SC 3α-diol, 3β-diol, androsterone, or oil vehicle, and tested in the same tasks. Experiment 3: GDX rats were androsterone- or vehicle-primed and administered an antagonist of ARs (flutamide), ERs (tamoxifen), or GBRs (flumazenil), or vehicle and then tested in the elevated plus maze. Both rats and wildtype mice, but not βERKO mice, consistently had reduced anxiety and improved performance in the object recognition task. Androsterone was only effective at reducing anxiety-like behavior in the elevated plus maze and this effect was modestly reduced by flumazenil administration. Thus, actions at ERβ may be required for T's anxiety-reducing and cognitive-enhancing effects.     

Interaction of nutritional and socioeconomic status as determinants of cognitive development in disadvantaged urban Guatemalan children

The relationship between nutritional status, measured by height for age, and cognition, measured by WISC full-scale IQ, was studied in a longitudinal sample of 459 urban Guatemalan children, aged 4-9 years, from a disadvantage community of the fringe of Guatemala City, examined annually over a 3 year period. Socioeconomic status (SES) was controlled by developing a composite index for each home. The mean IQ differed significantly from the lowest to the highest quartiles of stature, the difference between the two extreme quartiles being approximately 0.3 SD of IQ. There was a significant interaction between SES and stature in their effects on IQ. Whereas nutritional status and SES affected IQ in an additive fashion in the upper three SES quartiles, there was no difference in IQ attributible to stature among children from the most disadvantaged homes. This analysis indicates that the mild-to-moderate protein-energy malnutrition (defined by height for age) that is prevalent among children from disadvantaged environments in developing countries is significantly related to cognitive development. However, in the poorest homes, SES is seen as a more important determinant of cognitive development than stature.     

The Influence of Internal Time,Time Awake,and Sleep Duration on Cognitive Performance in Shiftworkers

To date, studies investigating the consequences of shiftwork have predominantly focused on external (local) time. Here, we report the daily variation in cognitive performance in rotating shiftworkers under real-life conditions using the psychomotor vigilance test (PVT) and show that this function depends both on external and internal (biological) time. In addition to this high sensitivity of PVT performance to time-of-day, it has also been extensively applied in sleep deprivation protocols. We, therefore, also investigated the impact of shift-specific sleep duration and time awake on performance. In two separate field studies, 44 young workers (17 females, 27 males; age range 20–36 yrs) performed a PVT test every 2?h during each shift. We assessed chronotype by the MCTQ^Shift (Munich ChronoType Questionnaire for shiftworkers). Daily sleep logs over the 4-wk study period allowed for the extraction of shift-specific sleep duration and time awake in a given shift, as well as average sleep duration (“sleep need”). Median reaction times (RTs) significantly varied across shifts, depending on both Local Time and Internal Time. Variability of reaction times around the 24 h mean (≈ ±5%) was best explained by a regression model comprising both factors, Local Time and Internal Time (p < .001). Short (15th percentile; RT_15%) and long (85th percentile; RT_85%) reaction times were differentially affected by Internal Time and Local Time. During night shifts, only median RT and RT_85% were impaired by the duration of time workers had been awake (p?<?.01, consistent with the highest sleep pressure), but not RT_15%. Proportion of sleep before a test day (relative to sleep need) significantly affected median RT and RT_85% during morning shifts (p?<?.01). RT_15% was worst in the beginning of the morning shift, but improved to levels above average with increasing time awake (p < .05), whereas RT_85% became worse (p < .05). Hierarchical mixed models confirmed the importance of chronotype and sleep duration on cognitive performance in shiftworkers, whereas the effect of time awake requires further research. Our finding that both Local Time and Internal Time, in conjunction with shift-specific sleep behavior, strongly influence performance extends predictions derived from laboratory studies. (Author correspondence: roenneberg@lmu.de)