The vestibular system provides vital information about head position and motion; which is used for the control of balance through vestibulospinal reflexes. Chapter 2 explores the process of transforming head position to body coordinates, with and without vision. The results show when vision is available, the evoked response is less precise. Chapter 3 explores the transformation process before and after 60 days of bedrest. After this period of inactivity, participants swayed more, and their EVS-evoked sway response was less precise. This decrement in precision appears to begin recovery 6 days postbedrest. Chapter 4 focuses on vestibulo-ocular reflexes rather than postural reflexes. Electrical vestibular stimulation is used to evoke measurable torsional eye-movements. The magnitude of the response is modulated by stimulus frequency. Results suggest that CNS interprets electrical vestibular stimulation as a velocity signal rather than a position or acceleration signal. This technique is an ideal measure of pure vestibular function, Chapter 5 utilised the technique in a clinical environment. Vestibular schwannoma patients, with known unilateral vestibular deficit, were tested to identify if the proposed technique can detect this deficit. Results showed that asymmetries could be detected, and, the test may be more sensitive than previously used measures of vestibular asymmetries.

The human fetal adrenal cortex is highly active, producing large amounts of Δ5 androgens. Together with the placenta, the fetal adrenal cortex regulates circulating progesterone, estrogen and corticosteroids in the fetus and has a major effect on maternal steroid levels. At birth, production of adequate aldosterone and cortisol are essential for survival, and shortly after birth in the human the cortex undergoes rapid remodelling. Catecholamines, produced by the adrenal medulla are also vital to the body’s ability to respond to stress, and regulate metabolic processes and blood pressure. Normal development of both the cortex and medulla of the human fetal adrenal are therefore critical for post-natal human endocrinology and health. Despite this, adrenal development and function during fetal life is poorly understood due both to the inherent difficulties in obtaining and studying human samples and to the unique nature of human adrenal/endocrine developmental which means animal models are of limited value. While poorly understood, the processes involved in fetal development are highly regulated although they can be disrupted by exposure to environmental chemicals or endocrine disruptors. Exposure to maternal smoking, for example, can disrupt normal fetal programming and has been linked with an altered postnatal stress response in the offspring, as well as a higher risk of developing cardiovascular disease, diabetes, and metabolic syndrome in adult life. These programming effects point to the possible involvement of the human fetal adrenal gland in the post-natal pathologies associated with maternal smoking. The aims of the studies described here were to develop a method to measure 26 steroids by liquid chromatography mass spectrometry (LC/MS) from samples, from which RNA and protein were also extracted, in order to make optimal use of small amounts of human tissue and use this method, and others, to examine the development and steroidogenic capacity of the human fetal adrenal cortex as well as the migration and development of the pheochromoblast cells which will go on to form the adrenal medulla. The influence of maternal smoke-exposure was also examined in these tissues. Additional aims of a set of in vitro studies were to examine the effects of cigarette smoke extract (CSE) and its components on the steroidogenic capacity of a human pluripotent adrenocortical cell line, as well as the influence of placental steroids and their withdrawal. 109 human fetal adrenals were obtained from elective terminations (REC 04/S0802/21) of second trimester fetuses between 11-21 weeks of gestation. Fetuses were grouped according to sex, gestational age and maternal smoking. Cortical steroids extracted from these adrenals were quantified by LC/MS. Cortical and medulla development was examined by measurement of key elements of the steroidogenic, catecholaminergic, and angiogenic pathways using real-time PCR (RT-qPCR), western blot and immunohistochemistry. Statistical analysis of the data was carried out using generalised linear models with age, sex and maternal smoking status as covariates. For in vitro smoke-exposure experiments, H295R cells were cultured for 3 full days in the presence of cotinine, nicotine, or CSE, and stimulated with forskolin. For examinations of the effects of placental steroids, cells were cultured in the presence of estrogen and/or progesterone for 4 days followed by an additional 4 days of culture without added steroids. Steroids and mRNA transcript levels from cells were measured by ELISA, LC/MS and RT-qPCR. Data was analysed using mixed-effects general linear models. The most abundant steroid (ng/mg of tissue) in the human fetal adrenal was pregnenolone, followed by dehydroepiandrosterone-sulphate and 17-hydroxyprogesterone and levels of these steroids were similar between male and female fetuses. Cortisol was present in all adrenals examined although aldosterone was undetected. This data suggests adrenal regulation of circulating fetal ACTH remains a factor in development of congenital adrenal hyperplasia during the 2nd trimester while lack of aldosterone probably explains salt-wasting disorders frequently seen in extreme preterm neonates. Fetal plasma levels of ACTH, intra-adrenal levels of progesterone, 17α-hydroxyprogesterone and 16α-hydroxyprogesterone and adrenal transcript levels of the transcription factors GATA-6 and NR5A1 were increased by maternal smoking. However, plasma and intra-adrenal cortisol, and intra-adrenal DHEAS were unaffected. The enzymes involved in catecholamine biosynthesis were all highly expressed in the developing medulla. Migrating prechromoblast cells were clearly visible in the adrenal fetal zone through H&E staining and could be categorised into noradrenaline- and adrenaline-producing cells by immunohistochemistry for tyrosine hydroxylase and phenylethanolamine-N-methyl transferase. Maternal smoking was also associated with increased levels of PHOX2B transcript, a transcription factor involved in the maturation of chromaffin cells. In cell culture experiments, levels of CYP11A1, CYP17A1, CYP21A2, HSD3B, PGR and ESR2 transcripts were all significantly reduced in cells exposed to CSE. The effects of CSE-exposure on steroid production was variable but pregnenolone and progesterone production was highly stimulated under basal conditions, indicating that CSE has both cell-stimulatory and cell-inhibitory effects. Estrogen and progesterone exposure also had variable effects on steroid production by the cells, depending on other stimulatory factors, and results indicate that other placental factors, in addition to placental steroids, are likely to play a role in cortical remodelling after birth. In conclusion, the studies described here have comprehensively examined the steroidogenic capacity of the human fetal adrenal cortex during the second trimester and have begun to characterise the development and migration of the pheochromoblasts. The results described here provide an understanding of normal development of the steroidogenic capacity of the human fetal adrenal during the second trimester, as well as some of the pathologies associated with abnormal adrenal development. The effects of maternal smoking on the processes examined were not marked, however the disruption of steroid production or dysregulation of transcription factors may lead to changes in adrenal function in postnatal life.

Ovulation has been hypothesized as an inflammatory process. Interleukin(IL)-1$\alpha$, IL-1$\beta$ and tumor necrosis factor(TNF)-$\alpha$ are potent cytokines produced from macrophages and various other cell types, and are pivotal components of inflammation. Although previous studies have investigated cytokine activities in the reproductive system, there is little information on their precise localization and activities during the periovulatory period. To investigate the role of cytokines in ovulation, experiments were designed to determine the immunohistochemical localization and time specific production of cytokines IL-1 and TNF-$\alpha$ using a mouse model at 36h, 12h, 6h, 2h before ovulation, and at 6h and 18h after ovulation in vivo. Isolated individual follicles in vitro were used to determine more precise roles of cytokines on follicular development, ovulation and steroidogenesis. From these studies it was found that (1) granulosa cells were the primary sites of IL-1$\alpha$ and TNF-$\alpha$ production from large antral follicles and preovulatory follicles in vivo, (2) production of IL-1$\alpha$ and TNF-$\alpha$ increased as ovulation neared, first appearing in the cumulus cells and expanding to antral and mural granulosa cells, (3) less intense staining of these cytokines in the theca layer of smaller follicles suggests that theca cells may contribute to the production of these cytokines to some extent, (4) but there was no IL-1$\beta$ production, (5) localized and temporal production of cytokines during the periovulatory period suggests precise regulation, (6) decrease of IL-1$\alpha$ in the ovary after gonadotropin injection determined by enzyme linked immunoadsorbent assay suggests that IL-1$\alpha$ production may be under the control of gonadotropins, (7) in follicle culture without bone marrow derived cells, granulosa cells were confirmed as the main source of cytokine production, (8) addition of IL-1$\alpha$ and TNF-$\alpha$ to follicles in culture tend to decrease estradiol production. In conclusion, immunoreactive cytokine production correlated positively with the periovulatory follicular development suggesting their role as ovulatory mediators. It requires further studies on what are the signals for the initiation and termination of cytokine production, how transcription and translation of these cytokines are regulated during the periovulatory period, and how they contribute to the ovulation.

Blood pressure, heart rate, and renal sympathetic nerve responses were measured in 9-13 week old male spontaneously hypertensive rats (SHR) and compared to those in age and sex-matched Wistar-Kyoto (WKY) rats following intravenous injection of the neuropeptide substance P (SP), the nicotinic stimulant 1,1-dimethyl-4-phenylpiperazinium (DMPP), and the adrenoceptor stimulant norepinephrine (NE). Charles River Sprague-Dawley (CD) rats were used in some studies to develop methodologies. Measurements were made in control rats and also following sinoaortic denervation, pithing, ganglion blockade, or adrenoceptor blockade. Responses were evaluated in order to determine if ganglion stimulation by SP was enhanced in SHR compared to WKY rats and if this enhancement was selective for SP or would also be exhibited to DMPP. NE was used to evaluate adrenergic sensitivity and to confirm the success of baroreceptor denervations. SHR exhibited greater intrinsic sympathetic tone than WKY rats before and following ganglion blockade. Ganglion stimulation by SP and DMPP was only fully revealed following elimination of baroreceptor input. Results indicated that SP stimulates sympathetic ganglia to increase renal sympathetic nerve activity, heart rate and blood pressure in CD, SHR and WKY rats. This increase was enhanced in SHR compared to WKY rats in the absence of a similar enhancement of responses to DMPP. The action of SP to cause vasodilation was attenuated in SHR versus WKY rats which may augment its action as a pressor agent in SHR. In conclusion, increases in blood pressure, heart rate and renal sympathetic nerve activity were selectively increased to SP in SHR versus WKY rats. This enhanced action of SP may contribute to the elevation of basal and/or evoked sympathetic discharge observed in this model of hypertension.

The purpose of this study was to precisely locate, in living humans, a myofascial trigger point associated with the upper portion of the trapezius muscle (TrP1) that refers pain to the head and neck and to determine if this point is associated with anatomical structures. This study is descriptive and utilizes data from measurements of the location of TrP1 in relation to anatomical landmarks, of pressure sensitivity overlying the trigger point and electromyography recordings in localizing the trigger point. Information obtained from living humans was used to determine anatomical correlation to structures in cadavers. Results indicated there is little variability in the location of TrP1 among individuals or from one extremity to the other, and this point may be associated with structures of the skin. A neurovascular supply (NAV) emerging from the upper trapezius to the skin was located in cadavers resembling the location of TrP1 in living humans. This NAV contained only small diameter nociceptive nerve fibers. Conclusion from the study show that TrP1 in living humans can be precisely located and that the mechanism of pain referral may involve structures of the skin. Future studies to precisely locate other myofascial trigger points may aid in identifying mechanisms of trigger point activation as well as aid clinicians in more precisely locating trigger points for treatment.

A series of trials was carried out to examine the role of prolactin (PRL) in the control of lactogenesis in New Zealand Romney x Border Leicester ewes. In addition, a study was made of differences in milk yields and plasma PRL concentrations between spring- and autumn-lambing ewes.Daily subcutaneous injections of 2 mg CB154 inhibited PRL secretion and delayed lactogenesis. There were no consistent effects on plasma progesterone or insulin concentrations. CB154 treatment was more effective in reducing milk yield in twin-bearing than in single-bearing ewes when used for 20 days than for 9 days prepartum. The differential effects on milk yield cannot be explained by corresponding effects on plasma PRL or insulin concentrations. Circulating PRL during the period 20 to 10 days prepartum may have an important effect on milk yield in twin- but not single-bearing ewes.Subcutaneous injections of 0.5 mg/kg live weight oPRL, administered on 2 consecutive days peripartum, to ewes treated with CB154 for 7 days prepartum, resulted in milk yields similar to those in control ewes and significantly (P<0.01) greater than those in ewes treated with CB154 alone. This indicated that oPRL prevented the CB154-induced reduction of milk yields and has established that the effect of CB154 on lactogenesis is mediated through suppression of PRL secretion and not by effects on some other hormone.Injection of 10 mg oPRL directly into one mammary gland (via the teat duct) increased milk yields relative to the contralateral, bicarbonate-treated gland in CB154-treated ewes. The intramammary oPRL injection did not raise circulating PRL concentrations. Furthermore, the milk yields of bicarbonate-treated glands in ewes treated with bicarbonate only, did not differ from those of bicarbonate-treated glands in ewes treated with oPRL in the contralateral gland, demonstrating that there were no effects of oPRL, transferred via the circulation from the treated gland, on the contralateral gland. Glands treated with oPRL produced 15% (P<0.05) more milk than the bicarbonate-treated glands during the first 8 days of lactation and the difference was maintained throughout the 8-week lactation period, indicating that the oPRL had effected a permanent change in the ability of the gland to produce milk. It is concluded that PRL acts directly on the mammary gland without the need for a putative intermediate hormone, and that intramammary PRL concentrations during lactogenesis may have long-lasting effects on lactation.The possibility was examined that dietary differences were responsible for seasonal differences in plasma PRL concentrations, milk yields, milk composition, lamb birthweight and lamb growth rate, observed in earlier trials. Mean plasma PRL levels were significantly (P<0.01) higher in spring- (192±38 ng/ml) than in autumn- (71±17 ng/ml) lambing ewes housed indoors under constant photoperiod (18L:6D) and fed the same diet. Milk yields were also significantly (P<0.05) higher in the spring- (2041±114 g/d) than in the autumn- (1563±109 g/d) lambing ewes over the 8 day lactation. Lamb growth rates (adjusted for birthweight, birthrank and sex of lamb) from birth to 8 weeks of age were significantly (P<0.001) higher in spring (282±12 g/d) than in autumn (225±15 g/d). The seasonal differences were confounded with corresponding differences in ewe live weight and it was not possible to determine whether dietary differences contributed significantly to the differences observed.Two routes of oPRL supplementation were used to test the effectiveness of elevating peripheral or local levels of PRL in autumn-lambing ewes which, based on previous results, were expected to have low plasma PRL concentrations and milk yields relative to spring-lambing ewes. Administration of 10 mg supplementary oPRL directly into the gland or subcutaneous injection of 0.5 mg/kg oPRL did not increase the milk yields, or change the composition of milk, compared to controls. These results suggest that the circulating level of PRL, and the intramammary concentration of PRL, in autumn-lambing ewes are not limiting lactogenesis. Because the plasma prolactin concentration in the ewes was unexpectedly high, it was not possible to reach firm conclusions regarding possible effects of supplementary oPRL in ewes with naturally low plasma PRL concentrations. Nevertheless, the results indicate that raising the intramammary concentration of PRL around the time of parturition, in ewes with circulating PRL levels characteristic of normal spring-lambing ewes, does not enhance lactogenesis.It is concluded that PRL is important to the complete initiation of lactogenesis in ewes, that it acts directly on the gland and that it is necessary for establishing the maximum potential of the gland to secrete milk.

One the most serious health hazards of aging and of long term space flight is the loss of bone. The most important determinant of the debilitation due to bone loss is the peak bone mass achieved during late adolescence, which itself may be influenced by gender and environmental factors such as mineral balance. Sodium intake is considered a risk factor for both hypertension and osteoporosis. It is estimated that 30-60% of the population is hypertensive and 30-40% of the population is salt sensitive. The primary purpose of the present studies was to both delineate and combine the effects between salt intake and salt sensitive hypertensive genotype on bone. Our hypothesis was that hypertensive rats would have more severely affected bone than normotensive rats due to salt supplementation and/or genotype. In addition, how these effects might be altered by immobilization/overloading stress was examined as this further burdens NASA space pioneers. This study investigated the possible effects of an ad libidum 1% or 2% saline instead of water on the normotensive (W) and salt sensitive hypertensive (SS) young female rats. A total of 46 weight-matched female rats (7 weeks old) were used. Treated rats in the 1% study drank 1% saline ad libitum for a 42 day salt supplementation period, beginning at day 7 (after arrival) to day 49. Treated rats in the 2% study drank 2% saline ad libitum for a 42 day salt supplementation period, beginning at day 7 (after arrival) to day 49. The right hindlimb of each animal was immobilized by binding to the abdomen with 4 layers of elastic bandage tape, the hip joint in flexion and the knee and ankle joint in extension for the 42 day salt experimental period. Body weight and urine volume was measured biweekly. Food and fluid intake was monitored daily. After sacrifice, three sites (both the underloaded and the overloaded tibiae, as well as the L-2 vertebrae) were processed for histomorphometric analysis. The wet weight (g) and length (mm) of the excised right immobilized and left overloaded femur and the ulna were measured. A 3-point bending test was applied to femurs only. Immediately after the femur breaking strength measurements, bone was cut transversely, one mm from breaking point (fracture location), and a 1.0 mm cross-section was cut for morphological measurement. In addition, a 5 mm high cylinder section from each femur was cut and used for bone composition measurements along with the right ulna bone. A number of elements were analyzed at one time with Induced Coupled Plasma (ICP) spectrometry. The systolic blood pressure and heart rate were measured in the 6th week of study by the tail-cuff sphygmomanometer method. A more robust result was seen with increased concentration of saline treatment from a 1% saline threshold level, to the 2% saline level. Using two-way ANOVA, both hypertensive genotype and 2% saline treatment significantly increased blood pressure and heart rate, and decreased femur magnesium. The SS rat had significant reductions in bone mass, femur cross-sectional area and zinc concentrations with simultaneous elevations in femur stiffness, strength and calcium concentrations. Two percent saline treatment markedly increased both blood pressure and heart rate and decreased both femurs magnesium and cancellous bone in the weight-bearing tibia bone. After 6 weeks of immobilization (to simulate space weightlessness), reductions in cancellous tibia bone volume, with elevations in femur bone stiffness, mineral concentration (calcium and phosphorus) and in trace elements (zinc and manganese) were found in the underloaded femur. Our findings suggest genotype, and saline treatment, and immobilization adversely affect bone in adolescent female rats. In addition, the deleterious bone effects are site specific, affecting each site differently.

The maintenance of function in an ageing population is essential to ensure current and future health in older people. The ability to walk independently in a range of situations and environments is key to successful ageing. Age-related gait adaptations including spatial-temporal parameters, joint kinematics and kinetics have been identified to be a consequence of the ageing process. For example, reduced walking speed and increased pelvic tilt are suggestive of compensation strategies to minimise falls. The majority of research has compared young adults (20-40 yrs) to older adults (≥ 50 yrs), categorising older adults into a single group regardless of actual age. An alternative approach is to explore the effects of age on gait and functional movement characteristics within an older adult population. One-hundred and fifty-eight community-dwelling older adults, age range 55 to 86 years (65.7 ± 6.8 yrs) were recruited to create a new gait database. Three-dimensional motion analysis captured five walking tasks: normal walking (with and without force plate contact), manual dual task walking and walking with obstacle clearance (stepping onto, off and over an obstacle). Age-related adaptations to walking occurred from age 75 years by adopting a joint kinetic strategy (including reduced hip extension moment) and altering gait (including a reduced walking speed). Increasing the task complexity was associated with altered gait patterns for this older adult group including a reduction in toe-clearance during manual dual task walking (increasing the likelihood of tripping) and increased arm swing during obstacle clearance (potentially increasing stability). This work represents the creation of one of the largest databases of gait in older people including three-dimensional motion analysis for normal walking and three functional walking tasks for healthy high-functioning older adults. It has the potential to be used to identify factors that predispose older adults to falling or with previously unidentified pathological changes.

Nonmetric traits (NMTs) are often used by osteoarchaeologists in the study of human variation. Some NMTs are affected by environmental factors whereas others are genetic in origin. Such genetic variants have long been used to support the hypotheses on the history and divergence of human populations suggesting that some population groups can be genetically distinguished. However, when genetic NMTs occur in higher than expected frequency these can be interpreted as possible indicators of relatedness. This method is applied to a sample of 977 individuals from the Medieval Poulton Chapel, St. Owen’s Church and Norton Priory Collections, U.K. One hundred and twenty-six cranial and postcranial NMTs were examined to determine: 1) the prevalence, 2) whether there are significant differences between the sexes and/or by age category, 3) if there is variation in mechanical and genetic NMT frequency between the three samples and, 4) to explore possible familial relationships through hierarchal cluster analysis and burial spatial distribution. It is thought that family members are often buried near one another, suggesting that individuals sharing similar genetic NMTs would be buried within close proximity to each other. This thesis has revealed the frequency of 126 NMTs for each sample. No significant differences were reported between the sexes at Poulton Chapel whereas significant differences were noted at St. Owen’s Church and Norton Priory, especially for NMTs considered genetic in origin. For all samples, significant differences were found among the age categories. Intra-populational differences were explored between the three samples. The results of these comparisons highlight that 60 NMTs are shared between the Poulton Chapel and Norton Priory Collection, while St. Owen’s Church only shares few traits with both sites. This suggests a probable geographical north-south divide between the three sites. Finally, the hierarchal cluster analysis identified probable familial relationships for the Norton Priory sample. This is supported by the burial spatial distribution and historical documentary evidence. Unfortunately, this analysis was unsuccessful for St. Owen’s Church with limited results for the Poulton Chapel sample. Future research is required to incorporate aDNA analysis to confirm the likelihood of familial links within these sites, supporting the use of certain NMTs is the use of establishing familial relationships.

Within the genus Homo, we observe a decrease in mandibular robusticity and in the size of anterior and postcanine dentition, a trend that is usually referred to as reduction or gracilisation. Factors linked to diet, food processing and encephalization have been suggested to be the main drivers of this trend. Stone tools and fire would have allowed Pleistocene hominins to reduce food toughness, thus relaxing the selective pressures on the masticatory apparatus. In the Holocene, the changes in human lifestyle triggered by agriculture would have determined the reduction in human tooth size. Brain expansion may have acted as a constraint on the development of the lower jaw. In this work, a primate perspective was adopted to clarify the relative influence of adaptive and non-adaptive factors on mandibular and dental reduction in the genus Homo. The effect of diet and structural constraints (allometry and encephalization) on dental and mandibular size and robusticity were analysed. The results show that incisor size and mandibular robusticity correlate significantly with diet proxies in non-human extant catarrhines and with neurocranium shape changes in the neurocranium in Homo sapiens. In non-human African apes, the elongation of the neurocranium influences postcanine tooth size. In Homo, body size plays an important part in tooth size allometry, but not in robusticity. These results suggest that improvements in tool-based food preparation may have been a leading factor in the reduction of incisor size in hominins. Molars and premolars were probably influenced by the expansion of the neurocranium during Pleistocene, and incisor size may be constrained by neurocranium shape changes in H. sapiens. This work confirmed the importance of food processing in the trend of reduction and produced convincing evidence for the significance of structural constraints in the evolution of the hominin anatomy. These findings contribute to explain the complex evolution of the human skull.