Calcium and magnesium physiology and nutrition in relation to the prevention of milk fever and tetany (dietary management of macrominerals in preventing disease).

Dairy cows may suffer events of hypocalcemia and hypomagnesemia, commonly known as milk fever and tetany. Milk fever is characterized by hypocalcemia at parturition as a consequence of a sudden increase in Ca demand and an unavoidable delay in Ca metabolism adaptation. Tetany is due to impaired Mg absorption from the rumen that cannot be compensated by absorptive or excretory adaptation, resulting in a net nutritional shortage of Mg and culminating in hypomagnesemia. Prevention strategies require triggering the activation of Ca gastrointestinal absorption and avoiding factors limiting ruminal Mg absorption.

[Electrophysiological analysis of functional state of spinal cord motoneurons in rats with parathyroprivous tetany].

In normal rats and of those with parathyroprivous (hypocalcemic) tetany the comparative analysis of background activity (BA), tetanic and posttetanic increase and decrease of frequency of spinal cord (SC) motoneurons (MNs) responses under high-frequency (50, 100Hz) stimulation (HFS) of flexor (G) and extensor (P) hind-limb nerves have been conducted. The on-line selection and program analysis of the spikes was produced. On the 3-7 and 21-22 days of development of acute and chronic tetany, respectively, the significant tetanic and posttetanic changes of MNs activity without meaningful changes in BA was registered. Along with the abrupt increase of excitatory manifestation of activity to HFS in a period of development of acute tetany was observed their relative weakening in animals with chronic tetany. Simultaneously the weakening or total disappearance of depressor reaction, especially expressed in the period of development of acute tetany was noted. It was concluded on the causal dependence of the parathyroprivous convulsions due to disturbances of correlation of inhibitory-excitatory processes in SC MNs.

IP(3)-dependent, post-tetanic calcium transients induced by electrostimulation of adult skeletal muscle fibers.

Tetanic electrical stimulation induces two separate calcium signals in rat skeletal myotubes, a fast one, dependent on Cav 1.1 or dihydropyridine receptors (DHPRs) and ryanodine receptors and related to contraction, and a slow signal, dependent on DHPR and inositol trisphosphate receptors (IP(3)Rs) and related to transcriptional events. We searched for slow calcium signals in adult muscle fibers using isolated adult flexor digitorum brevis fibers from 5-7-wk-old mice, loaded with fluo-3. When stimulated with trains of 0.3-ms pulses at various frequencies, cells responded with a fast calcium signal associated with muscle contraction, followed by a slower signal similar to one previously described in cultured myotubes. Nifedipine inhibited the slow signal more effectively than the fast one, suggesting a role for DHPR in its onset. The IP(3)R inhibitors Xestospongin B or C (5 µM) also inhibited it. The amplitude of post-tetanic calcium transients depends on both tetanus frequency and duration, having a maximum at 10-20 Hz. At this stimulation frequency, an increase of the slow isoform of troponin I mRNA was detected, while the fast isoform of this gene was inhibited. All three IP(3)R isoforms were present in adult muscle. IP(3)R-1 was differentially expressed in different types of muscle fibers, being higher in a subset of fast-type fibers. Interestingly, isolated fibers from the slow soleus muscle did not reveal the slow calcium signal induced by electrical stimulus. These results support the idea that IP(3)R-dependent slow calcium signals may be characteristic of distinct types of muscle fibers and may participate in the activation of specific transcriptional programs of slow and fast phenotype.

Overexpression of HSP10 in skeletal muscle of transgenic mice prevents the age-related fall in maximum tetanic force generation and muscle Cross-Sectional Area.

Skeletal muscle atrophy and weakness are major contributors to frailty and impact significantly on quality of life of older people. Muscle aging is characterized by a loss of maximum tetanic force (P(o)) generation, primarily due to muscle atrophy, to which mitochondrial dysfunction is hypothesized to contribute. We hypothesized that lifelong overexpression of the mitochondrial heat shock protein (HSP) HSP10 in muscle of mice would protect against development of these deficits. P(o) generation by extensor digitorum longus muscles of adult and old wild-type and HSP10-overexpressing mice was determined in situ. Muscles were subjected to damaging lengthening contractions, and force generation was remeasured at 3 h or 28 days to examine susceptibility to, and recovery from, damage, respectively. Muscles of old wild-type mice had a 23% deficit in P(o) generation and a 10% deficit in muscle cross-sectional area compared with muscles of adult wild-type mice. Overexpression of HSP10 prevented this age-related fall in P(o) generation and reduction in cross-sectional area observed in muscles of old wild-type mice. Additionally, overexpression of HSP10 protected against contraction-induced damage independent of age but did not improve recovery if damage occurred. Preservation of muscle force generation and CSA by HSP10 overexpression was associated with protection against the age-related accumulation of protein carbonyls. Data demonstrate that development of age-related muscle weakness may not be inevitable and show, for the first time, that lifelong overexpression of an HSP prevents the age-related loss of P(o) generation. These findings support the hypothesis that mitochondrial dysfunction is involved in the development of age-related muscle deficits.

When EGF is offside, magnesium is wasted.

Our understanding of magnesium (Mg(2+)) regulation has recently been catapulted forward by the discovery of several disease loci for monogenic disorders of Mg(2+) homeostasis. In this issue of the JCI, Groenestege et al. report that their study of a rare inherited Mg(2+) wasting disorder in consanguineous kindred shows that EGF acts as an autocrine/paracrine magnesiotropic hormone (see the related article beginning on page 2260). EGF stimulates Mg(2+) reabsorption in the renal distal convoluted tubule (DCT) via engagement of its receptor on the basolateral membrane of DCT cells and activation of the Mg(2+) channel TRPM6 (transient receptor potential cation channel, subfamily M, member 6) in the apical membrane. These authors show that a point mutation in pro-EGF retains EGF secretion to the apical but not the basolateral membrane, disrupting this cascade and causing renal Mg(2+) wasting. This work is another seminal example of the power of the study of monogenic disorders in the quest to understand human physiology.

Impaired basolateral sorting of pro-EGF causes isolated recessive renal hypomagnesemia.

Primary hypomagnesemia constitutes a rare heterogeneous group of disorders characterized by renal or intestinal magnesium (Mg(2+)) wasting resulting in generally shared symptoms of Mg(2+) depletion, such as tetany and generalized convulsions, and often including associated disturbances in calcium excretion. However, most of the genes involved in the physiology of Mg(2+) handling are unknown. Through the discovery of a mutation in the EGF gene in isolated autosomal recessive renal hypomagnesemia, we have, for what we believe is the first time, identified a magnesiotropic hormone crucial for total body Mg(2+) balance. The mutation leads to impaired basolateral sorting of pro-EGF. As a consequence, the renal EGFR is inadequately stimulated, resulting in insufficient activation of the epithelial Mg(2+) channel TRPM6 (transient receptor potential cation channel, subfamily M, member 6) and thereby Mg(2+) loss. Furthermore, we show that colorectal cancer patients treated with cetuximab, an antagonist of the EGFR, develop hypomagnesemia, emphasizing the significance of EGF in maintaining Mg(2+) balance.

Transverse tubular system depolarization reduces tetanic force in rat skeletal muscle fibers by impairing action potential repriming.

When muscle fibers are repeatedly stimulated, they may become depolarized and force output decline. Excitation of the transverse tubular system (T-system) is critical for activation, but its role in muscle fatigue is poorly understood. Here, mechanically skinned fibers from rat fast-twitch muscle were used, because the sarcolemma is absent but the T-system retains normal excitability and its properties can be studied in isolation. The T-system membrane was fully polarized by bathing the skinned fiber in an internal solution with 126 mM K(+) (control solution) or set at partially depolarized levels (approximately -63 and -58 mV) in solutions with 66 or 55 mM K(+), respectively, and action potentials (APs) were triggered in the sealed T-system by field stimulation. Prolonged depolarization of the T-system reduced tetanic force proportionately more than twitch force, with greater effect at higher stimulation frequency (responses at 20 and 100 Hz reduced to 71 and 62% in 66 mM K(+) and to 54 and 35% in 55 mM K(+), respectively). Double-pulse stimulation showed that depolarization increased the repriming period (estimated minimum time before a second AP can be produced) from approximately 4 ms to approximately 7.5 and 15 ms in the 66 and 55 mM K(+) solutions, respectively. These results demonstrate that T-system depolarization reduces tetanic force by impairing AP repriming, rather than by preventing AP generation per se or by inactivating the T-system voltage sensors. The findings also explain why it is advantageous to reduce the rate of motoneuron stimulation to muscles during repeated or prolonged periods of activity.

Hypomagnesaemia and new data on vitreous humour magnesium concentration as a post-mortem marker in ruminants.

Magnesium deficit has been associated with many sub-clinical and clinical conditions in humans and animals. The incidence of hypomagnesaemia is high in lactating cows grazing spring pastures, occasionally resulting in the often fatal condition known as grass tetany. While plasma magnesium concentrations can be used to assess magnesium status in the live animal, post-mortem diagnosis of clinical grass tetany is difficult. Recent studies have investigated the potential of eye fluid magnesium concentration as a post-mortem marker of hypomagnesaemic tetany. In tetany induction studies carried out in adult ewes and lactating cows significant relationships were found to exist between the concentrations of magnesium in either cerebrospinal fluid or plasma and either aqueous or vitreous humour. In freshly dead animals aqueous humour magnesium concentrations of < 0.33 mmol/L in adult sheep and < 0.25 mmol/L in adult cattle were associated with severe hypomagnesaemia and tetany. However, aqueous humour was found to be unstable post-mortem. Vitreous humour was considerably more stable and a vitreous humour magnesium concentration in adult sheep of < 0.65 mmol/L for up to 24 hours post-mortem or < 0.55 mmol/L in adult cows for up to 48 hours was associated with severe hypomagnesaemia and tetany. Provided clear-fluid samples are taken from appropriate animals and processed correctly in the laboratory, the concentration of magnesium in vitreous humour is a useful and practical marker in the post-mortem diagnosis of hypomagnesaemic tetany in ruminants.

Oxidant production and immune response after stretch injury in skeletal muscle.

PURPOSE: This study investigated oxidant production and associated immune response after acute muscle stretch injury. METHODS: A standardized single stretch injury was performed on the tibialis anterior (TA) muscle of 36 male New Zealand white rabbits while contralateral control limbs underwent a sham surgery. Animals were sacrificed 0, 4, 12, 24, 48, and 72 h after injury. Potential sites of oxidant production, measured with a dichlorofluorescein (DCF) probe, were evaluated using two separate buffers. RESULTS: Nonmitochondrial oxidant production measured under basal buffer conditions (0.1 M potassium phosphate) was increased in both injured and control limbs at 24 h (P < 0.01) and was greater in the injured limb at 12 and 48 h (P < 0.01). There was also an interaction of time and injury (P < 0.05). Maximum oxidant production by neutrophils and macrophages, stimulated by the induced buffer (including 1.7 mM ADP, 0.1 mM NADPH, 0.1 mM FeCl3), was increased in both injured and control limbs at 4 h (P < 0.01) and was greater in the injured limb at 48 h (P < 0.01). Myeloperoxidase (MPO) activity, indicating the presence of activated neutrophils, was higher in the injured limb at 4 and 48 h (P < 0.01). The activities of superoxide radical producing and quenching enzymes, xanthine oxidase (XO) and superoxide dismutase (SOD), were elevated at 24 (P < 0.01) and 4 h (P < 0.05), respectively, but showed no difference between injured and control limbs. CONCLUSION: We conclude that acute muscle stretch injury and the required surgeries to generate the injury result in a biphasic increase in oxidant production in both injured and control limbs, suggesting a systemic immune response. The increase in oxidant production at 4 h may be caused by an increase in activated neutrophils, whereas XO activity may contribute to oxidant generation at 24 h.

Regional brain monoamine concentrations and their alterations in bovine hypomagnesaemic tetany experimentally induced by a magnesium-deficient diet.

Monoamines are important brain neurotransmitters. An investigation was carried out to determine if hypomagnesaemic tetany was associated with alterations in regional brain monoamine concentrations in bovines. The results, established in cows with normal magnesium status, demonstrated that regional differences existed in the distribution and concentration of brain monoamines in the adult bovine, which were similar to those in other species. In magnesium-deficient cows, severe hypomagnesaemia and lowered cerebrospinal fluid (CSF) magnesium concentrations were associated with significant alterations in monoamine concentrations in some brain regions. Alterations in 3,4-dihydroxyphenylalanine (DOPA) and dihydroxyphenylacetic acid (DOPAC) concentrations in the corpus striatum, and dopamine (DA) in the cerebral cortex and cerebellum were recorded. These regions play an important role in both voluntary and involuntary motor function, and therefore these alterations may play a role in the aetiology of hypomagnesaemic tetany. However, there was no significant change in DA concentrations in the corpus striatum (the main dopaminergic region in the brain) associated with hypomagnesaemia. In addition, a significantly lower norepinephrine (NE) concentration in the corpus striatum of hypomagnesaemic animals was also recorded. Norephinephrine is generally excitatory and therefore lowered NE concentrations would be expected to result in depression rather than stimulation of motor function.

[Tetany--a first symptom of celiac disease]. / Tetani--debutsymptom ved cøliaki.

BACKGROUND: Seizures in children are usually caused by fever or epilepsy, though they may also be caused by electrolyte, vitamin or mineral disturbances. We describe a case of hypocalcaemic tetany with hypovitaminosis D. MATERIAL AND METHODS: The patient was a previously healthy eight months old girl of Indian origin. RESULTS: Investigations revealed that she suffered from coeliac disease. INTERPRETATION: Coeliac disease is a well-known disorder, characterised by enteropathy and malabsorption causing symptoms such as vomiting, diarrhoea and failure to thrive. Enteropathy and specific malnutrition may occur in the total absence of gastrointestinal symptoms.

Counter effects of higenamine and coryneine, components of aconite root, on acetylcholine release from motor nerve terminal in mice.

The counter effects of higenamine and coryneine, components of aconite root, on acetylcholine (ACh) release from motor nerve terminals in the mouse phrenic nerve-diaphragm muscle preparation were studied by a radioisotope method. Both nerve-evoked release and spontaneous release of [3H]-ACh from the preparation preloaded with [3H]-choline were measured. The change in the tetanic tension of muscle was simultaneously recorded in the same preparation. Higenamine (10 microM) augmented both the nerve-evoked and spontaneous ACh releases, and the muscle tension. The effects were inhibited by pretreatment with propranolol (10 microM), a beta-adrenoceptor antagonist. Coryneine reduced the nerve-evoked release of ACh, accelerated the decay of tetanic tension (tetanic fade) at 30 microM, and it depressed the peak amplitude of tetanic tension at a higher concentration of 100 microM. These results suggest that of the two components contained in aconite root, higenamine increases ACh release via activation of beta-adrenoceptor, and conversely coryneine depresses ACh release by preferentially acting at motor nerve terminal.

Effects of nitric oxide on the contraction of skeletal muscle.

A review of the literature suggests that the effects of nitric oxide (NO) on skeletal muscles fibers can be classified in two groups. In the first, the effects of NO are direct, due to nitrosation or metal nitrosylation of target proteins: depression of isometric force, shortening velocity of loaded or unloaded contractions, glycolysis and mitochondrial respiration. The effect on calcium release channels varies, being inhibitory at low and stimulatory at high NO concentrations. The general consequence of the direct effects of NO is to 'brake' the contraction and its associated metabolism. In the second group, the effects of NO are mediated by cGMP: increase of the shortening velocity of loaded or unloaded contractions, maximal mechanical power, initial rate of force development, frequency of tetanic fusion, glucose uptake, glycolysis and mitochondrial respiration; decreases of half relaxation time of tetanus and twitch, twitch time-to-peak, force maintained during unfused tetanus and of stimulus-associated calcium release. There is negligible effect on maximal force of isometric twitch and tetanus. The general consequence of cGMP-mediated effects of NO is to improve mechanical and metabolic muscle power, similar to a transformation of slow-twitch to fast-twitch muscle, an effect that we may summarize as a 'slow-to-fast' shift.

[Magnesium balance in patients with spasmophilia. Relation to results of electromyography]. / Magneziová bilance u nemocných se spazmofilií. Vztah k výsledkum elektromyografie.

BACKGROUND: The pathophysiological basis of spasmophilia is frequently magnesium deficiency and the therapeutic administration of magnesium salts has usually a favourable effect. However the parameters of magnesium balance are not always consistent with the results of electromyography. The objective of the present work was to test and interpret the relationship of results of these two basic diagnostic procedures indicated when spasmophilia is suspected. METHODS AND RESULTS: Thirty-three subjects (9 men and 24 women) with suspected spasmophilia were examined by non-invasive electromyography, using the technique of surface electrodes. All subjects had concurrently biochemical examinations: serum calcium and ionized calcium, serum magnesium (S-mg), magnesium in erythrocytes (ery-Mg) and magnesium in the blood haemolysate (H-Mg). In 29 patients and oral magnesium loading test was made with evaluation of the urinary Mg excretion after a constant Mg load (U-Mg). Statistical evaluation of the investigated parameters of the magnesium balance revealed a highly significant relationship between ery-Mg and U-Mg and H-Mg and ery-Mg (p < 0.005). A less close relationship was found between H-Mg and S-Mg (p < 0.05). Total and ionized calcium was in all examined subjects within the range of the arbitrary normal range. The EMG finding was positive (the finding of two and more multiplets in the ischaemic and hyperventilation test resp.) in 30 instances, i.e. in 91% of the examined subjects. In 72% there was agreement of the positivity of the EMG and magnesium deficiency (i.e. reduced values of ery-Mg and U-Mg), positivity of EMG combined with normal parameters of the Mg balance was recorded in 18%. In 6.1% of the examined subjects magnesium deficiency was confirmed combined with a normal EMG finding. CONCLUSIONS: Concurrent positivity of EMG and magnesium deficiency in 72% justifies the therapeutic administration of magnesium. In patients with a normal magnesium deficiency and positive EMG another cause of spasmophilia must be taken into consideration, incl. technical errors of interpretation of EMG results. A negative EMG associated with magnesium deficiency can suggest the central form of tetany, where magnesium treatment is also unequivocally indicated.

The effect of muscle length on intracellular calcium and force in single fibres from mouse skeletal muscle.

1. The effect of muscle length on the myoplasmic free Ca2+ concentration ([Ca2+]i) and tetanic force production of intact single muscle fibres was investigated in the mouse flexor brevis muscle. Muscle fibres were analysed at 100 microns intervals from 100 microns shorter than the optimum length (L(zero)) to 300 microns longer than L(zero). The fluorescent Ca2+ indicator indo-1 was used to measure [Ca2+]i. 2. Changes in muscle length did not significantly affect tetanic or resting [Ca2+]i. However, at lower stimulus frequencies (30-50 Hz) the force-length relation was shifted to longer muscle lengths. 3. Force-tetanic [Ca2+]i curves were constructed at each muscle length. A comparison of these curves revealed that the Ca2+ sensitivity was length dependent. Ca2+ sensitivity increased over the ascending limb and plateau phase of the force-length curve, but over the descending limb Ca2+ sensitivity reached a plateau and then started to decrease once more. 4. These results show that length-dependent changes in Ca2+ sensitivity of single muscle fibres cannot simply be related to variations in sarcomere length. It is proposed that other factors, such as cross-bridge attachment and/or developed force, have a role in determining the Ca2+ sensitivity at a particular muscle length.

[Electrolyte changes during and after voluntary hyperventilation]. / Elektrolytveränderungen während und nach willkürlicher Hyperventilation.

Paresthesia and tetanic finger cramps during hyperventilation-induced respiratory alkalosis are believed to derive from a pH-dependent decrease of ionized serum calcium. In the study reported here, ionized serum calcium, total calcium and total protein were measured during a three-minute hyperventilation period in ten volunteers. During hyperventilation finger paresthesias appeared in all probands without proof of any significant change in ionized serum calcium (1.26 +/- 0.05 mmol/l at the end of the three-minute hyperventilation period). Total protein increased as a consequence of hyperventilation-induced transient hemo-concentration. Paresthesias and tetanic finger cramps during the three-minute hyperventilation could not be related to changes of ionized serum calcium; however the other electrolytes, i.e. sodium, magnesium, potassium, chloride, phosphate and bicarbonate, showed, with the exception of sodium, significant changes.

Neurotrophic factor mRNA expression in dentate gyrus is increased following in vivo stimulation of the angular bundle.

Nerve growth factor (NGF) and brain-derived neurotrophic factor (BDNF) are two structurally-related neurotrophins synthesized in dentate gyrus granule cells and pyramidal neurons of the hippocampal formation. These neurons receive excitatory glutamatergic afferents from the entorhinal cortex via the angular bundle/perforant path. In the present study, we tested whether electrophysiological stimulation of this glutamatergic pathway modifies NGF or BDNF messenger RNA (mRNA) expression in vivo. Within hours following brief trains of high frequency angular bundle stimulation, the levels of mRNA encoding both neurotrophins were increased exclusively in granule cells of the ipsilateral dentate gyrus. The increase in neurotrophic factor mRNA expression was found to be mediated through the N-methyl-D-aspartate (NMDA) glutamate receptor subtype, and occurred in the absence of seizure. These findings provide evidence that neurotrophic factor mRNA levels in the hippocampal formation are increased by direct activation of excitatory afferents originating in the entorhinal cortex. We suggest that the function of some neurotrophin-responsive neuronal populations may depend upon the integrity and activity of neurons in the entorhinal cortex, a population of neurons reported to be compromised in patients with Alzheimer's disease.

Calcium concentration changes in the calyciform nerve terminal of the avian ciliary ganglion after tetanic stimulation.

A study has been made of the changes in calcium concentration in the calyciform nerve terminal ([Ca]c) and in the neurone soma ([Ca]s) of avian ciliary ganglion cells following tetanic stimulation of the nerve terminal. Dissociated ciliary neurones were loaded with the calcium indicator Fura-2 and digital imaging techniques used to determine the spatial and temporal distribution of calcium in the cells during post-tetanic potentiation (PTP) and long-term potentiation (LTP). Stimulation of the calyciform terminal with an extracellular electrode at 10 Hz for 2 s increased both [Ca]s and [Ca]s over 3-fold, with the [Ca] increasing for each impulse in the facilitatory train. The increase in [Ca]s could be prevented by allowing the terminal to degenerate in culture before stimulation. Stimulation of the calyciform terminal with a long tetanus of 30 Hz for 20 s gave an over 4-fold increase in both [Ca]c and [Ca]s by the end of the train. Analysis of the decline in [Ca]c after the train showed that it disappeared from the calyx along a double exponential time course with time constants of about 1 min and 50 min, respectively. These times are similar to those of PTP and LTP in the ganglia, and are almost independent of the extracellular calcium level. In order to determine whether the influx of calcium ions during a tetanus was through N-type calcium channels, these were blocked with adenosine (100 microM). Adenosine blocked the increase in both [Ca]s and [Ca]c that normally accompanies a tetanus. Thapsigargin (200 nM) did not affect [Ca]c or [Ca]s, but blocked transient increases in [Ca] caused by caffeine (10 mM) in both 3 mM and Ca2+ free bath solutions. These results are discussed in relation to the role of intracellular calcium in initiating LTP after a tetanus to the nerve terminals.