NMDA receptors (NMDARs) are heteromeric assemblies of NR1 and NR2(A-D) subunits with properties heavily influenced by the type of NR2 subunit incorporated. While NMDARs with only one type of NR2 subunit have been extensively characterized, little is known about receptors containing two different NR2 subunits, despite compelling evidence that such triheteromeric receptors exist in vivo. We used a point-mutation approach that allows isolation of recombinant triheteromeric NMDARs possessing two different NR2 N-terminal domains (NTDs). We show that in receptors associating the NR2A-NTD (sensing nanomolar Zn) and the NR2B-NTD (sensing ifenprodil), each NTD binding site retains selective high affinity for its ligand. However, each ligand produces only partial inhibition, and maximal inhibition requires occupancy of both NR2-NTDs by their respective ligands. Similarly, NR1/2A/2C receptors are inhibited by zinc with high potency but low efficacy. Therefore, interactions between homologous N-terminal domains determine the unique pharmacological properties of triheteromeric NMDARs.

Properties of maximum likelihood estimators of rate constants for channel mechanisms are investigated, to see what can and cannot be inferred from experimental results. The implementation of the HJCFIT method is described; it maximises the likelihood of an entire sequence of apparent open and shut times, with the rate constants in a specified reaction mechanism as free parameters. The exact method for missed brief events is used. Several methods for testing the quality of the fit are described. The distributions of rate constants, and correlations between them, are investigated by doing sets of 1000 fits to simulated experiments. In a standard nicotinic receptor mechanism, all nine free rate constants can be estimated even from one single channel recording, as long as the two binding sites are independent, even when the number of channels in the patch is not known. The estimates of rate constants that apply to diliganded channels are robust; good estimates can be obtained even with erroneous assumptions (e.g. about the value of a fixed rate constant or the independence of sites). Rate constants that require distinction between the two sites are less robust, and require that an EC50 be specified, or that records at two concentrations be fitted simultaneously. Despite the complexity of the problem, it appears that there exist two solutions with very similar likelihoods, as in the simplest case. The hazards that result from this, and from the strong positive correlation between estimates of opening and shutting rates, are discussed.

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The mechanisms that underlie activation of nicotinic receptors are investigated using human recombinant receptors, both wild type and receptors that contain the slow channel myasthenic syndrome mutation, epsilonL221F. The method uses the program HJCFIT, which fits the rate constants in a specified mechanism directly to a sequence of observed open and shut times by maximising the likelihood of the sequence with exact correction for missed events. A mechanism with two different binding sites was used. The rate constants that apply to the diliganded receptor (opening, shutting and total dissociation rates) were estimated robustly, being insensitive to the exact assumptions made during fitting, as expected from simulation studies. They are sufficient to predict the main physiological properties of the receptors. The epsilonL221F mutation causes an approximately 4-fold reduction in dissociation rate from diliganded receptors, and a smaller increase in opening rate and mean open time. These are sufficient to explain the approximately 6-fold slowing of decay of miniature synaptic currents seen in patients. The distinction between the two binding sites was less robust, the estimates of rate constants being dependent to some extent on assumptions, e.g. whether an extra short-lived shut state was included or whether the EC50 was constrained. The results suggest that the two binding sites differ by roughly 10-fold in the affinity of the shut receptor for ACh in the wild type, and that in the epsilonL221F mutation the lower affinity is increased so the sites become more similar.

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A comprehensive experimental study has been made of the NM-64 neutron monitor. The counting rate has been measured as a function of the thickness of the reflector, of the separation of the counters, of the thickness of the inner moderator, and of the gas pressure in the counters. It is shown that the figures given in the internationally approved specification represent a satisfactory optimization of the design. The counting rate of the NM-64 monitor is found to be 3.3 times that of an IGY monitor of the same horizontal area. The origin and movement of both fast and thermal neutrons within the monitor have been investigated using cadmium sheets and by studying events producing counts in more than one counter.

Whole‐cell patch‐clamp recordings were used to investigate possible age‐related changes in K+ currents of type I carotid body cells isolated from the rat. K+ current density increased with age, as measured in cells isolated from 4‐day‐old, 10‐day‐old and adult rats (≥ 5 weeks old). The proportion of current reversibly inhibited by high [Mg2+] (6 mm), low [Ca2+] (0.1 mm) solutions, indicative of the proportion of current attributable to activation of Ca2+‐sensitive K+ (KCa) channels, was significantly smaller in cells of 4‐day‐old rats compared with 10‐dayold rats, despite inward Ca2+ current densities being similar in these two age groups. Inhibition of K+ currents by high [Mg2+], low [Ca2+] solutions was similar in 10‐day‐old and adult type I cells. Hypoxia (PO2, 16–23 mmHg) caused reversible reductions in type I cells from rats of all age groups. However, reductions seen in cells of 4‐day‐old rats were significantly smaller than those seen in cells of 10‐day‐olds and adults. The degree of hypoxic inhibition in these latter two groups was not significantly different. In the presence of high [Mg2+], low [Ca2+] solutions, hypoxia (PO2, 16–23 mmHg) was without significant effect on residual K+ currents in cells from all age groups. These observations indicate that K+ current density increases with postnatal age in the rat. Between days 4 and 10, there appears to be a predominant enhancement of KCa channels, and over the same age range hypoxic sensitivity of K+ currents increases. Our findings demonstrate that this latter observation arises because hypoxia selectively inhibits KCa channels in cells at all ages studied. These results suggest an important role for KCa channels in postnatal maturation of hypoxic chemoreception in the rat carotid body.

SCCMS mutations may show a recessive inheritance pattern and variable penetrance. A diagnosis of SCCMS should not be ruled out in cases of CMS with an apparent recessive inheritance pattern.