A system for accurate and automated injection of hyperpolarized substrate with minimal dead time and scalable volumes over a large range.

Over recent years hyperpolarization by dissolution dynamic nuclear polarization has become an established technique for studying metabolism in vivo in animal models. Temporal signal plots obtained from the injected metabolite and daughter products, e.g. pyruvate and lactate, can be fitted to compartmental models to estimate kinetic rate constants. Modeling and physiological parameter estimation can be made more robust by consistent and reproducible injections through automation. An injection system previously developed by us was limited in the injectable volume to between 0.6 and 2.4ml and injection was delayed due to a required syringe filling step. An improved MR-compatible injector system has been developed that measures the pH of injected substrate, uses flow control to reduce dead volume within the injection cannula and can be operated over a larger volume range. The delay time to injection has been minimized by removing the syringe filling step by use of a peristaltic pump. For 100µl to 10.000ml, the volume range typically used for mice to rabbits, the average delivered volume was 97.8% of the demand volume. The standard deviation of delivered volumes was 7µl for 100µl and 20µl for 10.000ml demand volumes (mean S.D. was 9 ul in this range). In three repeat injections through a fixed 0.96mm O.D. tube the coefficient of variation for the area under the curve was 2%. For in vivo injections of hyperpolarized pyruvate in tumor-bearing rats, signal was first detected in the input femoral vein cannula at 3-4s post-injection trigger signal and at 9-12s in tumor tissue. The pH of the injected pyruvate was 7.1±0.3 (mean±S.D., n=10). For small injection volumes, e.g. less than 100µl, the internal diameter of the tubing contained within the peristaltic pump could be reduced to improve accuracy. Larger injection volumes are limited only by the size of the receiving vessel connected to the pump.

Functional MRI in conscious rats using a chronically implanted surface coil.

PURPOSE: To establish procedures for functional MRI (fMRI) in rats without the need for anesthetic agents. MATERIALS AND METHODS: Rats were trained to habituate to restraint in a harness and scanner noise. Under anesthesia, rats were then prepared with a cranial implant that permitted stabilization of the head during subsequent imaging experiments. The cranial implant included an radiofrequency (RF) coil that was used to transmit and receive radiofrequency signals during imaging. Further training was then conducted to habituate the animals to head fixation whilst in the MR scanner. RESULTS: Using this method, we were able to successfully and repeatedly record BOLD fMRI responses to hypercapnia and whisker stimulation in awake rats. Electrical stimulation of the whisker pad produced a ∼7% increase in BOLD signal in the corresponding barrel cortex as well as adjacent negative BOLD responses, whilst hypercapnia produced larger increases in BOLD signal amplitude. CONCLUSION: This methodology leaves the face and limbs free from obstruction, making possible a range of behavioral or sensory stimulation protocols. Further development of this animal model could enable traditional behavioral neuroscience techniques to be combined with modern functional neuroimaging.

A novel task for the investigation of action acquisition.

We present a behavioural task designed for the investigation of how novel instrumental actions are discovered and learnt. The task consists of free movement with a manipulandum, during which the full range of possible movements can be explored by the participant and recorded. A subset of these movements, the 'target', is set to trigger a reinforcing signal. The task is to discover what movements of the manipulandum evoke the reinforcement signal. Targets can be defined in spatial, temporal, or kinematic terms, can be a combination of these aspects, or can represent the concatenation of actions into a larger gesture. The task allows the study of how the specific elements of behaviour which cause the reinforcing signal are identified, refined and stored by the participant. The task provides a paradigm where the exploratory motive drives learning and as such we view it as in the tradition of Thorndike [1]. Most importantly it allows for repeated measures, since when a novel action is acquired the criterion for triggering reinforcement can be changed requiring a new action to be discovered. Here, we present data using both humans and rats as subjects, showing that our task is easily scalable in difficulty, adaptable across species, and produces a rich set of behavioural measures offering new and valuable insight into the action learning process.

Neurovascular coupling is brain region-dependent.

Despite recent advances in alternative brain imaging technologies, functional magnetic resonance imaging (fMRI) remains the workhorse for both medical diagnosis and primary research. Indeed, the number of research articles that utilise fMRI have continued to rise unabated since its conception in 1991, despite the limitation that recorded signals originate from the cerebral vasculature rather than neural tissue. Consequently, understanding the relationship between brain activity and the resultant changes in metabolism and blood flow (neurovascular coupling) remains a vital area of research. In the past, technical constraints have restricted investigations of neurovascular coupling to cortical sites and have led to the assumption that coupling in non-cortical structures is the same as in the cortex, despite the lack of any evidence. The current study investigated neurovascular coupling in the rat using whole-brain blood oxygenation level-dependent (BOLD) fMRI and multi-channel electrophysiological recordings and measured the response to a sensory stimulus as it proceeded through brainstem, thalamic and cortical processing sites - the so-called whisker-to-barrel pathway. We found marked regional differences in the amplitude of BOLD activation in the pathway and non-linear neurovascular coupling relationships in non-cortical sites. The findings have important implications for studies that use functional brain imaging to investigate sub-cortical function and caution against the use of simple, linear mapping of imaging signals onto neural activity.

Deaf and hearing children: a comparison of peripheral vision development.

This study investigated peripheral vision (at least 30° eccentric to fixation) development in profoundly deaf children without cochlear implantation, and compared this to age-matched hearing controls as well as to deaf and hearing adult data. Deaf and hearing children between the ages of 5 and 15 years were assessed using a new, specifically paediatric designed method of static perimetry. The deaf group (N = 25) were 14 females and 11 males, mean age 9.92 years (range 5-15 years). The hearing group (N = 64) were 34 females, 30 males, mean age 9.13 years (range 5-15 years). All participants had good visual acuity in both eyes (< 0.200 LogMAR). Accuracy of detection and reaction time to briefly presented LED stimuli of three light intensities, at eccentricities between 30° and 85° were measured while fixation was maintained to a central target. The study found reduced peripheral vision in deaf children between 5 and 10 years of age. Deaf children (aged 5-10 years) showed slower reaction times to all stimuli and reduced ability to detect and accurately report dim stimuli in the far periphery. Deaf children performed equally to hearing children aged 11-12 years. Deaf adolescents aged 13-15 years demonstrated faster reaction times to all peripheral stimuli in comparison to hearing controls. Adolescent results were consistent with deaf and hearing adult performances wherein deaf adults also showed significantly faster reaction times than hearing controls. Peripheral vision performance on this task was found to reach adult-like levels of maturity in deaf and hearing children, both in reaction time and accuracy of detection at the age of 11-12 years.

Optical imaging spectroscopy in the unanaesthetised rat.

We describe a method for imaging the local cortical haemodynamic response to whisker stimulation in the rat without use of anaesthetic or paralytic agents. Female Hooded Lister rats were anaesthetised and a section of skull overlying somatosensory cortex thinned to translucency. A stainless steel chamber was then secured over the thin cranial window. Following recovery, animals were supported in a harness whilst the head was held by the implanted chamber using a pneumatically driven clamp. Optical imaging and optical imaging spectroscopy (OIS) of somatosensory cortex were performed whilst the contralateral whiskers were stimulated using a computer controlled air-puffer. Imaging sessions lasted approximately 15 min and data were collected for at least three consecutive days. Experiments were then repeated with the animals under urethane anaesthesia. Spectral analysis revealed qualitatively similar haemodynamic response functions across both anaesthetic states. However, our results indicate that the cortical haemodynamic response to somatosensory stimulation is larger by a factor of approximately 5 in the unanaesthetised rat compared with the anaesthetised rat. This preparation may make possible the investigation of the haemodynamic correlates of a broad range of neurological processes in the awake, behaving rodent.