Transactions of magnesium (Mg) along the gastrointestinal tract and the effect of change in potassium (K) intake were recorded in two in vivo experiments in sheep and cattle. Additional information on the sensitivity to K intake was obtained by comparing Mg transport and electrochemical properties of isolated rumen epithelia of sheep and cattle in 4 additional in vitro experiments. The experiment described in Chapter 2, and performed in sheep housed indoors in metabolic crates, investigated the compensatory capacity of the intestine to respond to the reduction in Mg absorption from the stomach as a consequence of increase in K intake. The animals were equipped with a ruminal cannula and two intestinal cannulae (duodenum and ileum) and flow of digesta was measured by the addition of two indigestible markers, chromium ethylenediaminetetra-acetic acid (Cr-EDTA) and ytterbium acetate (Yb). The animals were infused in a Latin square design for periods of 10 days with a solution of K bicarbonate that provided between 15 and 47 g of K/day. The diet consisted of a 50:50 combination of concentrates plus lucerne hay that provided around 3.7 g of Mg per day and 15 g of K per day. After 5 days of infusion samples of feed, faeces, urine and plasma were collected and analysed for Mg and K content. After 6 days of infusion, samples of duodenal and ileal flow were obtained. The treatments reproduced the detrimental effect of K on Mg absorption, especially in the rumen; a rise in K intake from 15 to 23 g/day reduced total Mg absorption from the gastrointestinal tract from 1.36 to 1.23 g/day, further increase in K intake to 38 and 47 g/day reduced absorption to 1.12 and 1.05 g/day, an overall reduction of around 50% in Mg apparent availability. Magnesium was mainly absorbed in the stomachs and large intestine with the small intestine a site of net secretion. Most of the reduction in Mg absorption with increase in K intake occurred in the stomachs, reducing from 1.86 to 1.11 g/day. A compensatory reduction in the net secretion of Mg from the intestines (small and large) was observed. This compensation was largely due to reduction in net secretion from the small intestine, from 0.85 to 0.22 g/day, rather than an increase in net absorption from the large intestine, although both segments acted synergistically. Results also suggested significant individual variation in plasma Mg concentration, urinary Mg excretion and in the flow and absorption of Mg along the gastrointestinal tract. It was suggested that most of that variability was due to genetic factors. Differences between species (cattle and sheep) were pursued during the course of the experiment described in Chapter three. Four triple cannulated rams and 3 triple cannulated dry cows were placed in metabolic crates, fed daily fresh-cut pasture and infused, in a total randomised design that provided the equivalent of an intake of 30,40 and 50 g of K per kg dry matter intake (DMI) per day. Solutions of K (as K bicarbonate) and markers (CrEDT A and Yb acetate) were infused continuously for a period of 10 days; after 5 days of infusion samples of pasture, faeces, urine and plasma were collected and analysed for Mg and K content. After 6 days of infusion, samples of duodenal and ileal flow were obtained. Total feed offered, refusals and water consumption were recorded daily. Results showed a greater sensitivity of cattle to the increase in K supply. A rise in K supply from 30 to 40 g per kg DMI/day reduced Mg absorption by almost 50% from 0.32 to 0.16 g per kg DMI/day, whereas only the highest treatment dose (50 g of K per kg DMI/day) produced the same deleterious effect in sheep. The absorption of Mg occurred mainly in the stomachs and large intestine; in contrast the small intestine was a site of net secretion in both species. The addition of K slightly reduced the rate of Mg absorption from the rumen, especially in cattle. Similarly, net Mg secretion within the intestines increased with increasing K intake in both species, only to be counterbalanced by a greater Mg absorption from the large intestine. The large intestine in both species (sheep and cattle) reduced faecal losses of Mg but was unable to fully compensate for the reduction in Mg absorption from the stomach or the greater net Mg secretion observed at the small intestine. Differences between species in water content of the faeces were observed to be mainly related to the moisture content of the digesta that reached the ileum rather than a result of differences in absorption in the large intestine. More evidence of species differences in Mg transport and of sensitivity to K intake were obtained by using isolated rumen epithelia and the Ussing chamber technique. Transport and electrophysiological properties of the tissues were observed in standard conditions and by adding different K concentrations to the mucosal side. Under standard conditions and open-circuit voltage, sheep isolated rumen epithelia had greater transmural potential difference (PDt), and lower conductance (Gt) but similar short-circuit current (Isc) than those from cattle. These results suggested that the rumen epithelium of cattle is leakier than that of sheep. Measurement of the transport of Mg showed that isolated rumen epithelia of cattle transported more Mg and was saturated at higher Mg concentrations (12 vs 4 mM) than sheep epithelia. These differences in Mg influx (transport from mucosa to serosa) were also observed in studies of Mg transport using stable isotopes. Magnesium influx (transport from mucosa to serosa) from the isolated rumen of cattle was greater than in sheep (57.5 ± 12.72 vs. 17.3 ± 12.72 nmol.cm⁻².h⁻¹); however this was counterbalanced by a greater Mg efflux (transport from serosa to mucosa) of 48.1 ± 12.72 vs. 9.9 ± 12.72 nmol.cm⁻².h⁻¹, for cattle and sheep respectively, when mucosal K concentrations were around 25 mM. A increase in K concentration on the mucosal side enhanced transmural potential difference (PDt) and short-circuit current (Isc) to a greater extent in sheep than in cattle, suggesting a greater effect of K on sheep than on cattle epithelia. On the other hand, the transport of Mg measured by stable isotopes suggested that net absorption of Mg (7.4 ± 12.72 vs. 11.1 ± 12.72 nmol.cm⁻².h⁻¹) in sheep epithelia was similar at 25 and 50 mM of K on the mucosal side, whereas net Mg influx in cattle was largely depressed as a consequence of a reduction in Mg influx (mucosa to serosa) from 57.7 ± 12.72 to 2.9 ± 12.72 nmol.cm⁻² h⁻¹ together with a constant Mg efflux (serosa to mucosa) 48.1 ± 12.72 and 41.2 ± 12.72 nmol.cm⁻².h⁻¹, presumably leaving through a paracellular shunt. However, this finding was based on date from a small size and caution should be applied to this conclusion. In conclusion, data collected from several comparative studies suggest differences in Mg apparent availability between sheep and cattle and also a greater sensitivity of cattle to an increase in K intake. This high sensitivity to K represents a great risk of hypomagnesaemia in dairy cattle in New Zealand where high K concentration is endemic in pastures. Most importantly, these results suggest that models for Mg metabolism in cattle should be based on measurements from cattle nutritional and physiological studies rather than on extrapolation from sheep studies.

Elk (Cervus elaphus spp) are widely used as a terminal sire in the New Zealand deer industry because elk red deer crosses are heavier at 12 months of age than pure-bred red deer (Cervus elaphus) and therefore better fit market demands. However, it is unclear whether nutritional requirements differ between genotypes. A series of experiments compared young (4 - 12 months) red deer and red deer-elk cross (hybrids) in various aspects of their nutrition. Single genotype groups (10-15) of red deer and hybrid weaner stags were offered one of four pasture allowances (2 to 12 kg DM/head/day) on a rotationally grazed mixed ryegrass - white clover pasture system for 9 weeks in winter (June-July), spring (October-December) and summer (February - March). Stags were weighed and given a new allocation of pasture weekly. Pre-grazing pasture mass ranged from 800 kg DM/ha for low pasture allowances to 4500kg DM/ha for higher allowances. Winter live-weight gain was low (40-80 g/day), relatively unaffected by pasture allowance and similar for both genotypes. In spring however, hybrids gained live-weight on average 100 g/day more than red deer across all pasture allowances and the response to additional pasture allowance was large (110 g/day at 2kg DM/head/day to 300 g/day at 9.5 kg DM/head/day). At the highest pasture allowance, hybrids grew faster (350 g/day) than red deer (250 g/day), although red deer were able to achieve this live-weight gain when offered less pasture (4 vs 12 kg DM/head/day, respectively). Summer live-weight gain was lower for both genotypes and responded less to increases in pasture allowance than during spring. A second experiment compared the live-weight gain of both genotypes at ad lib feeding in an indoor environment where intake could be accurately measured. A group of red deer (n =15) and a group of hybrid (n =15) weaner stags were housed indoors during winter (3 June - 27 August) and spring (16 October - 16 December) and fed a pelleted grain based ration ad lib. Mean daily intake for each group (kg DM/head/day) was calculated as the difference between feed offered and feed refused. Hybrids had a significantly higher (P< 0.05) absolute DM intake compared with red deer in both seasons, although when expressed on a metabolic body weight basis, there was no difference between genotypes irrespective of season. Live-weight gain during winter did not differ significantly between genotypes regardless of whether it was expressed on an absolute or metabolic weight basis. Spring live-weight gain, expressed both on an absolute and metabolic live-weight basis, was significantly higher for hybrids compared with red deer (P<0.05). Red deer and hybrids increased their feed intake from winter to spring by 20% and 24% respectively on a metabolic body weight basis. Although the difference between genotypes in their seasonal increase in intake was relatively small there was a large difference in their pattern of live-weight gain. Red deer exhibited a 34% and hybrids a 76% seasonal increase in live-weight gain expressed on a metabolic live-weight basis from winter to spring. These results indicate the greater rate of live-weight gain displayed by hybrids compared with red deer was not associated with a greater ad lib intake (expressed on a metabolic body weight basis) and the seasonal increase in live-weight gain is greater for hybrids than for red deer. A further experiment estimated the energy requirement for maintenance of both genotypes. Five deer of each genotype were housed in separate pens (3.5m²) during winter (3 June - 27 August) and spring (16 October - 16 December) and randomly assigned to one of 5 feeding levels (0.5, 0.6, 0.7, 0.8, or 0.9 times estimated ad lib intake of l.5 and l.7 kg DM/head/day during the winter and 3.0 and 3.3 kg DM/head/day during the spring for red deer and hybrids, respectively. Maintenance requirement was determined by regression analysis of live-weight gain on ME intake. Although there was no seasonal effect on the live weight gain response to intake there was a significant genotype effect. To maintain live weight during either season, hybrids required a higher ME intake (0.52 MJ ME/W0.75/day compared with red deer 0.41 MJ ME/W0.75/day). The rate of increase in live weight gain to increasing intake declined as intake increased and more so for red deer than hybrids. The final experiment in the series involved individually housed deer and aimed to more precisely determine differences in maintenance requirement and examine the difference in composition of gain between genotypes. In addition, in vivo apparent DM digestibility was measured in both genotypes. Red deer (n=7) and hybrid weaner stags (n=7) were housed in individual pens for a period of 8 weeks in both winter (July - August) and spring (November - December) and offered one of 7 feeding levels which ranged from maintenance to ad lib. During each 8 week experimental period, live weight gain, apparent digestibility and feed intake were measured. Immediately prior to, and at the conclusion of each 8 week period body composition was estimated using computer-assisted topography (CT scan). In winter, there was no significant difference in the live weight gain response to intake although red deer tended to have a higher (44 vs 55 MJ/kg) requirement for gain than hybrids. In spring, red deer had a lower requirement for maintenance (0.35 vs 0.47 MJ ME/W0.75/day) but a greater requirement for live weight gain (64 vs 35 MJ/kg) than hybrids. In spring, mean ad lib intake was about 30% higher than in winter and was greater for hybrids than for red deer. Energy retention in whole body (kJ/W0.75/day) did not differ between genotypes in either winter or spring but both the energy requirement for zero energy balance (0.59 vs 0.48 MJ ME/W0.75/day) and the efficiency of utilisation (0.37 vs 0.24) was greater in spring than in winter. The disparity between live weight gain and whole body weight gain may have been due to differences in gut fill. There was no significant difference between genotypes in relative growth coefficients for lean, bone or adipose tissue in whole body. However hybrids tended to have a higher winter and lower spring growth coefficient for fat compared with red deer. Growth coefficients for adipose, lean and bone, respectively were 0.983, 1.063 and 1.026 for winter and 1.02, 0.708 and 1.727 for spring. At the same whole body weight, deer in October had less adipose tissue than in August. It is unclear whether this represents a strategy for rapid spring growth or is an artefact of experimental protocol. Apparent dry matter digestibility (DMD) did not differ between genotypes but was higher by between 7 and 15 percentage units in winter compared with spring. Unexpectedly, digestibility was positively correlated with intake. Digestibility increased by 2.6 percentage units for every 10g DM/W0.75/day increase in either season in one group and 4.1 and 2.1 percentage units for deer in winter and spring respectively in another group. Errors in faecal collection were discounted as causes of the unexpected result.

Two experiments were conducted to examine the interrelationships between body composition at parturition, herbage allowance during lactation and milk production. Detailed measurements of feed intake and changes in body composition allowed estimates of maintenance energy requirement and efficiency of use of energy for milk production. Dorset ewes were offered low (L-) or high (H-) herbage allowances during pregnancy to achieve post-partum body weights, respectively, of about 50 and 60-65 kg. During the first 6 weeks of lactation herbage allowances were approximately 2 (-L), 5 (-M and -Mm) and 8 (-H) kg DM/ewe/d and feed intake was determined using chromic oxide dilution. In the first experiment (Expt.A) ewes were machine-milked and in the second (Expt.B) they were suckled by twin lambs with a further group (-Mm) machine-milked. Samples of ewes were slaughtered during early pregnancy, post-partum, and after 6 weeks of lactation. Body weight differences post-partum between L- and H- groups were 10 and 15 kg in Expts.A and B respectively. During lactation body weight changes appeared erratic, particularly in Expt.B, owing to the effects of variation in gut fill. Daily feed intake of machine-milked ewes on -M and -H allowances reached a maximum (1.9-2.2 kg OM/ewe/d) 2-3 weeks after parturition but ewes rearing lambs on similar allowances showed maximum intake (2.5-2.8 kg OM/ewe/d) during the first week. Mean intakes during lactation were 58, 69 and 73 g DOM/kg W•⁷⁵/d respectively for -L, -M and -H groups machine-milked and 51, 75 and 83 g DOM/kg W•⁷⁵/d respectively for -L, -M and -H groups rearing twin lambs. In both experiments ewes in L- groups had approximately 14% greater mean intake (g DOM/kg W•⁷⁵/d) during lactation compared with those in H- groups. Mean daily milk production of ewes rearing lambs was 2.0, 2.5 and 2.7 kg/d respectively in -L, -M and -H groups and was 33-52% greater than that for machine-milked ewes offered similar herbage allowances. In both experiments L- ewes had 14% greater milk yield (g/kg W•⁷⁵/d) compared with H- ewes. Mean lamb growth rates in Expt.B averaged 209, 254 and 268 g/lamb/d in -L, -M and -H groups, respectively. Lambs in the H- group were approximately 11% heavier at birth than those in the L- group and their mean growth rate was 20 g/d greater. Body fat and energy content of live ewes at the start of lactation was predicted using regression relationships from ewes slaughtered post-partum. There was large variation in the energy content of body weight loss during lactation (-37 to +140 MJ/kg) owing to variation in weight of gut fill and changes in chemical composition of the empty body. In Expt.A the body fat content of ewes during early pregnancy was 16.9 kg. Predicted values post-partum were 10.0 and 15.7 kg in L- and H- groups, respectively. During lactation all groups showed body fat losses which ranged from 53 (LH) to 120 g/d (HL). Body fat content during early pregnancy in Expt.B was slightly greater (19.0 kg) than in Expt.A and predicted values post-partum were 11.5 (L-) and 19.7 kg (H-). Fat mobilization during lactation was greater than in Expt.A, and ranged from 157 (LH) to 287 g/d (HL). In both experiments there were losses in body protein during pregnancy in L- ewes (5 and 15 g/d in Expts.A and B respectively). During lactation, protein losses were most evident in H- ewes, being greatest in HL groups (26 and 43 g/d in Expts.A and B respectively). Water: protein ratio in the empty body showed a progressive increase during pregnancy and lactation in both experiments. Estimated maintenance energy requirement for machine-milked ewes tended to be greater for H- (.236 MJ ME/kg W/d) compared with L- ewes (.205 MJ ME/kg W/d) but was similar for both groups of ewe rearing lambs (average of.238 MJ ME/kg W/d). Efficiency of use of ME above maintenance for milk production (K₁) in L- and H- groups was, respectively, .69 and .95 in machine-milked, and .69 and .64 in suckled ewes. Energy from mobilized body tissues was utilized for milk production with respective efficiencies in L- and H- ewes of .23 and .35 in machine-milked, and .40 and .50 in suckled ewes. Efficiency of conversion of total energy available (i.e. ME above maintenance and mobilized tissue energy) to milk energy (K₁(t)) when compared among all groups in both experiments, tended to decrease (from .84 to .51) with increasing body energy mobilization. There was a positive relationship, however, between K₁(t) and the proportion of mobilized energy derived from body protein. Metabolizable energy intakes in the present experiments appeared to be 10-15% greater than calculated requirements from ARC (1980). Estimates of ME requirements for lactating ewes at pasture, in relation to body weight, level of milk production and body energy change, have been calculated based on the present data.

Leucaena leucocephala and Sesbania sesban in dry form were fed as supplements to goats feeding on a low quality basal diet of barley straw. Each browse supplement was fed at four levels: 0 %, 0.83 % liveweight (LW), 1.66 % LW and at ad libitum. The intakes of dry matter (DM) and organic matter (OM), DM and OM digestibility, nitrogen (N) digestibility and balance, and liveweight gain were evaluated during a 5 week trial. For goats offered both leucaena and sesbania there was no significant (p<0.05) difference in DM or OM intakes between 0.83 % LW and 1.66 % LW levels of browse supplementation. When both lecaena and sesbania were offered ad libitum the DM and OM intakes were significantly (p<0.05) lower than for either 0.83 % LW or 1.66 % LW level of supplementation. Feeding both leucaena and sesbania increased the DM and OM digestibility coefficients significantly over thos of the controls. The growth rates for goats supplemented with leucaena at various levels were not significantly different from each other. For the goats offered sesbania at ad libitum, their growth rates were significantly lower than for those fed sesbania at either 0.83 % LW or 1.66 % LW. The apparent digestibility of nitrogen (ADN) was not significantly different among leucaena supplemented treatments. For goats offered sesbania, the ADN was significantly higher than for the controls. The ability of browse supplements to increase intakes and digestibility of both dry matter and organic matter is discussed.

Vitamin B₁₂ deficiency has been difficult to diagnose, mainly due to the vitamin's lack of biological significance in serum in which it is usually assayed. This research has investigated the marker of vitamin B₁₂/cobalt (Co) deficiency in sheep, methylmalonic acid (MMA), in comparison with serum and liver vitamin B₁₂ concentrations in farm situations where vitamin B₁₂ deficiency is expected in order to establish more accurate reference ranges for serum and liver vitamin B₁₂, and MMA. In addition, an attempt was made to ascertain the vitamin B₁₂ requirements of preruminant (PR) lambs, and to determine whether metabolic demand for vitamin B₁₂ influences tissue concentrations. Furthermore, since the vitamin is active in biological tissues in form of its coenzymes, 5’ -deoxyadenosylcobalamin and methylcobalamin, a preliminary assessment of variation in the distribution of these coenzymes in liver in different situations has been sought. The first trial was set up to find out if the addition of propionate to the PR lamb's diet stimulated the uptake and/or storage of vitamin B₁₂ in the liver as a reflection of the need to deal with the incoming propionate. Sixteen ten day old lambs (Dorset Down/Coopworth cross-bred) were housed indoors soon after birth and fed on milk replacer. For half of the lambs 7.5 % (w:w) of the milk powder was replaced by propionate. Within each group, four lambs were treated with 250 µg vitamin B₁₂ twice weekly. Supplementation with vitamin B₁₂ increased liver concentrations from ~250 to ~900 nmol/kg fresh tissue, but there was no effect of propionate. Propionate addition did, however, result in increased plasma vitamin B₁₂ concentrations in vitamin B₁₂ supplemented groups, values being 3323 and 2355 pmol/l in propionate supplemented and control groups, respectively. This suggested that diet could influence plasma vitamin B₁₂ concentrations. An attempt was made to quantify the PR lamb's ability to absorb vitamin B₁₂ from the alimentary tract by comparing the ability of intra-muscular (IM) and oral vitamin B₁₂ to raise plasma and liver vitamin B₁₂ concentrations. Twenty-seven three to four day old lambs from a farm with marginal Co status were housed indoors and fed on milk replacer. They were divided into three groups: control (n=3), IM treatment (n=12) and oral treatment (n=12). The two treatment groups were further subdivided into five sub-groups. These received, respectively, 0.2 (n=3), 0.4 (n=2), 0.8 (n=2), 1.6 (n=2) and 3.2 µg OH-cbl/d (n=3). The oral groups received tenfold the amount of the comparable IM groups, on the assumption that if oral absorption of the vitamin is about 10 % both groups would show similar increases in plasma and liver vitamin B₁₂ concentration. None of the IM groups showed any significant change in plasma or liver vitamin B₁₂. In the oral groups only the group on the highest dose of vitamin B₁₂, viz 32 µg/d, showed increases in plasma and liver concentrations. It was concluded that either absorption of vitamin B₁₂ was greater than 10 % or that the vitamin was retained better when administered orally. The amount retained in the livers of the lambs in the highest oral group was calculated to represent ~ 7.5 % of the dose. In a follow-up 24 h trial, 14 of the above lambs were divided into three groups: Control (n=3), oral (n=6) and IM (n=5) treatment. The IM group received 3.2 µg OH-cbl and the oral group tenfold the amount as single doses at 0800 h. Blood samples were taken at regular intervals throughout the 24 h period and assayed for vitamin B₁₂, Vitamin B₁₂ concentrations in the IM group rose steeply within the first hour after injection to a concentration that was calculated to reflect 100 % uptake of the vitamin. It rose more slowly over about 8 h in the oral group. From the area under the curve absorption of the oral dose was estimated to be ~ 7 %. The next experiment involved a farm where Co deficiency had been reported previously. In the first year, 50 pregnant two-tooth Half-bred ewes were divided randomly into two groups of 25. One group received a Co bullet plus 1000 µg OH-cb1 IM, the other group remained unsupplemented. In the following year the trial was repeated. Ewes from the previous year's trial (by then four-tooths) were augmented by a new cohort of pregnant two-tooths to make up numbers to 75. After lambing the lambs were divided into four groups: first by their dams' vitamin B₁₂ treatment, then half of each group received injections of vitamin B₁₂ at approximately three weekly intervals while the other half remained untreated. The trials lasted about five months, from mid-pregnancy until weaning. Pasture Co was at its lowest at lambing in both years, 0.09 and 0.10µg/g DM, respectively. In the first year, vitamin B₁₂ concentrations in the untreated ewes rose from 340 to 950 pmol/l in plasma and decreased in liver from 330 to 170 nmol/kg fresh tissue. In the Co treated group, vitamin B₁₂ concentrations in plasma rose from 500 to 1550 pmol/l and in liver from 310 to 560 nmol/kg fresh tissue. In the second year, vitamin B₁₂ concentrations in serum in the unsupplemented groups fell from 500 to 260 pmol/l around lambing before rising again to starting values at weaning, and liver vitamin B₁₂ concentrations fell from 450 at the start to 230 nmol/kg fresh tissue at the end of the trial. Serum vitamin B₁₂ concentrations in the two-tooth supplemented group rose from < 500 to > 3000 pmol/l whereas in the four-tooth supplemented group serum vitamin B₁₂ levels started at ~2800 and rose to nearly 5000 pmol/l. The supplemented four-tooths maintained higher liver vitamin B₁₂ concentrations throughout compared to the supplemented two-tooths, viz 680 compared to below 400 at the start and 900 versus 650 nmol/kg fresh tissue at weaning, respectively. MMA in the untreated groups rose to 15 and to 8 µmol/l during early lactation in the first and second years, respectively, whereas MMA in the treated groups stayed below 3 µmol/l in the first season and below 1.5 µmol/l in the second season. There was a live weight response to treatment in the ewes as the unsupplemented groups showed a significantly lower weight gain during the trials than the supplemented groups, viz 10.0 versus 13.6 kg in the first year, and 10.6 versus 13.3 kg in the four-tooths and 9.9 versus 12.1 kg in the two-tooths in the second year. There was also a significant difference in faecal egg count (FEC) in the first year. FEC in the untreated group was higher during lactation than in the treated group, viz 590 versus 170 eggs per gram wet faeces (epg), respectively. In the second year, the two-tooths had a higher FEC than the four-tooths, viz 120 versus 40 epg during the same time span, respectively. While there was a trend for treatment having an effect on FEC similar to that in the first year it was not significant. Supplementation of ewes in the first year increased mean milk vitamin B₁₂ concentrations at lambing from 800 to 1400 pmol/l and at weaning from 1750 to 4000 pmol/l. In the second year, Co bullet treatment increased milk vitamin B₁₂ concentrations in the four-tooths and two-tooths from 1500 and 2300 to 4000 and 2900 pmol/l at lambing, and from 1800 and 1400 to 6200 and 4500 pmol/l at weaning, respectively. Treatment of ewes increased vitamin B₁₂ concentrations in the lambs which were not themselves supplemented. Plasma values in the first year increased from 160 to 325 pmol/l soon after birth and from 650 to 900 pmol/l at weaning, and liver values from 75 to 140 nmol/kg fresh tissue soon after birth and from 150 to 240 nmol/kg fresh tissue at weaning. In the second year, plasma vitamin B₁₂ concentrations increased from 160 to 380 pmol/l soon after birth and from 500 to 700 pmol/l at weaning, and in liver from 130 to 260 nmol/kg fresh tissue soon after birth and from 220 to 340 nmol/kg fresh tissue at weaning. There was also a significant effect of ewe supplementation on lamb MMA in 1997/1998 when values decreased from 19 to 8 µmol/l around the time of rumen development. MMA in the second year stayed below 3 µmol/l throughout in all groups of lambs. There was no difference in LWG between any groups of lambs. FEC was lowest in the group where both ewes and lambs were supplemented and highest in the group where neither ewes nor lambs were treated. Further investigations were conducted on farms in Southland with lambs post-weaning in order to compare changes in serum and liver vitamin B₁₂ with serum MMA and LWG to determine the critical time and level of deficiency. In the first year, three farms with 50 lambs each participated. Lambs from each farm were allocated to five groups of 10 animals each. The first group received a Co bullet at weaning, and each month another group was treated with a Co bullet. The lambs were weighed monthly, and blood and liver samples were taken prior to treatment and each subsequent month from five lambs of the first supplemented group. The trial lasted about four months. Serum vitamin B₁₂ concentrations in lambs at weaning were between 500 and 1000 pmol/l. Although supplementation increased serum levels for the first month this was followed by a drop to near or below starting concentrations. An exception was Farm 3 where serum vitamin B₁₂ concentrations rose again at the end of the trial. Liver vitamin B₁₂ concentrations also showed an overall decline from starting levels (200 to 300 nmol/kg fresh tissue) to the end of the trial (100 to 200 nmol/kg fresh tissue). MMA started around 2 µmol/l and reached between 6 and 7 µmol/l in the untreated lambs on Farms 1 and 3 two months after weaning before decreasing to around 3 µmol/l at the end of the trial, whereas the treated lambs maintained MMA concentrations around 2 µmol/l. On Farm 2 MMA started just below 5 µmol/l, decreased to around 1 µmol/l for treated and untreated lambs one month later and rose again to between 2.5 and 4 µmol/l, respectively, at the end of the trial. LWG was below average for all lambs (between 0.20 and 0.04 kg/d except for Farm I in the first month after weaning) but no significant differences were noted between treated and untreated lambs on any of the farms. Another trial was conducted on one of these farms in the following year. One hundred lambs were divided into two groups of 50 each at weaning and sampled monthly for about six months. One group was treated with two Co bullets, the other group remained untreated. Pasture Co was between 0.04 and 0.07 µg/g DM, yet serum levels for the untreated group stayed ~500 pmol/l throughout the trial. Serum vitamin B₁₂ concentrations for the treated group started at ~500 pmol/l, rose to ~2500 pmol/l before falling back to ~2000 pmol/l. Liver vitamin B₁₂ concentrations for the untreated and treated groups were 529 and 427 nmol/kg fresh tissue at weaning, respectively. This decreased for both groups to ~350 nmol/kg fresh tissue one month after weaning. In the untreated lambs liver values decreased further to ~290 nmol/kg fresh tissue whereas they increased to ~450 nmol/kg fresh tissue for the treated group at the end of the trial. MMA concentrations started between 2 and 3 µmol/l for both groups and increased to 4.5 µmol/l for the untreated group one month later before falling back to 3.2 µmol/l. In the treated group MMA decreased to ~1µmol/l and stayed at that level throughout the trial. There was no difference in weight gain. In order to obtain an understanding of the distribution of corrinoids in biological tissues a High Performance Liquid Chromatography method was developed. The sensitivity of the analytical method meant that it was only practical to assay mainly liver samples because of the higher vitamin B₁₂ concentrations than in other tissues. The general finding was that the coenzyme 5’ –deoxyadenosylcobalamin (ado-cbl) constituted the highest proportion of corrinoids in liver (45 %), followed by analogues (28 %), OH-cbl (24 %) and lastly methy1cobalamin (3 %). Ado-cbl did tend to be proportionately higher in supplemented than in unsupplemented animals (56 and 42 %, respectively), whereas biologically non-active analogues tended to be higher in untreated than in treated sheep (29 and 21 %, respectively). It was concluded that in the farm trials Co deficiency was only mild or not present although deficiency would have been predicted from the low vitamin B₁₂ concentrations in serum and liver and from raised MMA values. Therefore, currently used thresholds in New Zealand appear to be too high for vitamin B₁₂, and overseas values for MMA do not seem to be appropriate. Revised marginal ranges of 100 to 200 pmol/l for serum, 100 to 200 nmol/kg fresh tissue for liver and 10 to 20 µmol/l for MMA are suggested. Further, this work shows that Co bullets were effective in elevating blood and liver vitamin B₁₂concentrations for longer than one year. In the trials with preruminant lambs it was found that maintenance requirements were met by the vitamin B₁₂ content of milk replacer. There is evidence from indoor and farm trials that vitamin B₁₂ from milk was much more readily absorbed than vitamin B₁₂ from supplements. It was estimated that suckling lambs probably require between 1200 and 4000 pmol vitamin B₁₂/d, depending on age.

A reduction in voluntary feed intake is a major factor in the lost productivity of grazing lambs infected by gastrointestinal parasites yet the mechanisms involved are poorly understood. Potential pathways involved in parasite-induced feed intake depression were investigated in lambs with minimal previous exposure to parasites and artificially infected by the small intestinal parasite Trichostrongylus colubriformis. Six in vivo experiments were conducted on lambs housed in individual pens or metabolism crates with similar feeding and experimental procedures. In Experiment 1 (Chapter 4) the effect of T. colubriformis infection on short term feed intake in lambs and of some pharmacological agents on feed intake depression were investigated. Prior to and for the duration of infection, lambs were fed once per day and feed intake recorded at regular intervals over the day (8 h). Following the onset of feed intake depression in the infected group (9 weeks after commencing dosing), all animals were treated with an analgesic (codeine phosphate per os), an anti-inflammatory agent (indomethacin per os), a CCK antagonist (L364-718 by subcutaneous injection) or saline (control) in a replicated Latin square design (n = 8). Although the pattern of feed consumption was similar in infected and non-infected lambs, average daily intake was reduced 32 % and short term intake (recorded at 10 minute intervals for the first hour of feeding, 15 minute intervals for the second hour and hourly for the next 6 hours of feeding) reduced 40 % by infection. This identified the key component by which intake was depressed and enabled the use of a short term intake model and short duration of action compounds to identify the pathways involved in intake depression in this sequence of experiments. None of the pharmacological treatments increased intake in the infected group. These results suggest a reduction in the rate of consumption due to reduced hunger signals, rather than change of meal eating patterns, is the major cause of feed intake depression. Specific conclusions about the pathways investigated using the pharmacological agents could not be obtained. Experiment 2 (Chapter 5) was designed to investigate the roles of pain and osmolality on feed intake depression. Digesta samples collected prior to and during parasite infection and before and after feeding had similar osmolalities (240-260 mosmol/l) which indicated that feeding or infection had no effect on osmolality of digesta. Following the onset of feed intake depression in infected animals, all animals were treated in a Latin square design (n = 4) with no treatment, saline, local anaesthetic (xylocaine) or analgesic (codeine phosphate) solution 15 minutes before feeding, by slow injection into the duodenum. There was no effect of these treatments on food intake. In the second part of the experiment, hyperosmotic solutions (mannitol and NaCI) markedly depressed short term intake in non-infected animals, suggesting a role for osmoreceptors in intake regulation. However these effects were not blocked by local anaesthetic so the depressed intake may have resulted from generalised malaise rather than from specific osmoreceptor effects. In Experiment 3 (Chapter 6) the role of peripheral CCK on intake depression was examined by a dose-response study utilising the CCK antagonist, loxiglumide. Intravenous injection of 5, 10 or 20 mg/kg LW of loxiglumide to infected lambs 10-15 minutes before feeding (n = 6) had no effect on feed intake at any of the dose levels. In experiment 4 (Chapter 7) loxiglumide was infused intravenously for 10 minutes (30 mg/kg/h) before feeding and for the first 2 h (10 mg/kg/h) after feed was offered to minimise any effect of the rate of clearance of loxiglumide on the lack of feed intake response. As well, the rate of marker disappearance from the abomasum was recorded in both infected and non-infected animals. Continuous infusion of loxiglumide did not attenuate parasite induced intake depression nor did it have any effect on abomasal emptying. Abomasal volume was reduced by infection (66.3 vs 162 ml) as was the fractional outflow rate (2.2 vs 2.8 ml/min) but these differences were accounted for by the lower level of feed intake in the infected animals. In Experiment 5 (Chapter 8) brotizolam, a benzodiazepine appetite stimulant, thought to act on the hypothalamus, was administered in a dose-response study to infected and non-infected animals (n = 4) immediately prior to feeding or following termination of the first meal (45 minutes after feeding) and the feed intake response recorded. Brotizolam elevated both the short term (0-0.75 h), daily (22 h) intake and all time intervals in the first 5 h after feeding in infected and non-infected animals when administered after the first meal but when administered prior to feeding elevated intake only over the first 6 h of feeding. In both cases the magnitude of the response was greater in infected animals than in non-infected animals. Brotizolam appeared to increase the rate of eating without having a major impact on meal eating patterns when administered before feeding. Where administration was after the first meal, the effect was due to an "extra" meal being consumed. These findings showed that infected animals can respond to central stimulators of intake although the mechanism of the response is not known. Opioids were implicated in intake depression as the rate of intake rather than meal patterns appeared to be the major parameter depressed under parasitism. This was examined in experiment 6 (Chapter 9) where animals (n = 6) were fasted for 26 h or not fasted, then treated with saline (control), brotizolam (intake stimulant) or naloxone (opioid antagonist) immediately prior to feeding. Fasting stimulated feed intake in the short term (100 % increase in 75 min) and over the day (12 % increase) in both infected and non-infected animals. Following fasting, infected animals ate a similar amount of feed to the non-infected, fasted animals and more than the non-infected, non-fasted animals. The signals resulting from a one day fast were sufficient in the short term to override parasite induced mechanisms causing feed intake depression. Naloxone suppressed the intake stimulatory effects of a 26 h fast in both infected and non-infected animals, which supports a role for endogenous opioids as hunger signals. Where animals were not fasted, naloxone reduced intake only in the non-infected animals which suggested endogenous opioid levels may be lower in infected animals than in non-infected animals. In the final experiment (Experiment 7, Chapter 10) the role of central hunger and satiety mechanism were investigated. Infected and non-infected animals (n = 6) were treated with naloxone or saline by intravenous injection, or saline and met-enkephalinamide (an opioid analogue) by intracerebral infusion, or naloxone and the opioid analogue simultaneously to investigate the role of central opioids in feed intake depression. To determine the role of CCK induced satiety signals on feed intake at a central level, loxiglumide and CCK were infused separately and in combination for 30 minutes prior to feeding and for the first 60 minutes of feed on offer, into a lateral cerebral ventricle of the brain of infected and control animals (n = 6). The opioid analogue tended to increase intake in infected animals but the effect was not significant probably because the dose used was too low to elicit a response in sheep. Naloxone depressed intake only in the infected animals, which conflicted with the results of Experiment 4. As a consequence these results were inconclusive because of the single low dose of opioid analogue used and the conflicting naloxone responses. CCK alone depressed intake by 39-52 % only in infected animals and this effect of the 90 minute infusion was evident over the 8 h short term recording period. Loxiglumide attenuated the feed intake depressive effects of CCK in the infected animals to the extent that intake was elevated above control levels. Loxiglumide alone was an intake stimulant in both infected and non-infected animals. Intake was increased over the entire 8 h but mostly in the second hour when intake was increased by 188 % in infected animals and by 16 % in the non-infected animals and resulted in almost continuous eating. These results showed loxiglumide will temporarily block the effect of parasite infection on feed intake in sheep when administered centrally and the fact that it blocked the effects of exogenous CCK on intake indicated that the effect is mediated via CCK receptors. In conclusion GIT parasite infection reduced both short term and daily feed intake apparently by a change in rate of intake rather than any alteration in meal patterns. It was further suggested that anyone of a number of potential peripheral pathways, including changes to osmolality, gut emptying, pain and inflammation of the gut, alone is not involved in anorexia in sofar as the compounds used could block these factors and the results support the idea that intake depression is mediated via a central mechanism. Intake in infected animals responded to a much greater extent when fasting, i.c.v. loxiglumide or brotizolam were employed. Feed intake thus appears to be regulated through the same mechanisms in infected and non-infected animals. The results from compounds affecting the central mechanism suggest central CCK receptors are important in parasite induced anorexia, possibly by changing the onset of satiety or by interacting with endogenous opioids to reduce the rate of feed intake. Secondly reduced endogenous opioids may be causing the reduction in the rate of feed consumption alone or as a result of other interactions. It was concluded that intake in parasitised animals could be increased to that of control animals by employing procedures and compounds thought to act on the hypothalamus.