Lactate dehydrogenase isoenzymes in sympathetic neurons and satellite gliocytes in normal conditions and in blockade of nicotinic cholinoreceptors.

Integral cytophotometry was used to measure lactate dehydrogenase (LDH) and its H-and M-isoforms in neurons and satellite gliocytes in tissue sections from the cranial cervical sympathetic ganglion of the rabbit in normal conditions and after experimental partial and complete pharmacological blockade of nicotinic cholinoreceptors (n-CR). In normal conditions, both cell types showed both the H-and M-type isoforms, though the isoenzyme profiles differed--neurons showed a dominance of H-isoform activity while the M-isoform was more active in satellite gliocytes. In partial and complete blockade, the activity of LDH and its H-and M-isoforms decreased significantly in proportion to the number of blocked n-CR. In satellite gliocytes, increases in the extent of blockade were associated with decreases in the activity only of the M-isoform, while the activity of the H-isoform did not change. In partial blockade, the LDH isoenzyme profile of satellite gliocytes shifted towards the neuronal isoform, while in complete blockade there was no difference from the LDH isoenzyme profile of intact neurons. These data led to the suggestion that the formation of lactate in satellite gliocytes is induced by nicotinic cholinergic synapses directly involved in neuron-glial interactions and in controlling the activity of the LDH enzyme system in sympathetic neurons.

Involvement of N-cholinergic synapses in regulating RNA metabolism in sympathetic neurons and satellite gliocytes.

RNA contents in the cytoplasm of neurons and satellite gliocytes in the cranial cervical ganglion of rabbits were determined by photographic cytophotometry during pharmacological blockade of N-cholinergic (N-Ch) synapses with the N-cholinolytic dimecoline (s.c. doses of 10, 30, and 50 mg/kg) and during the gradual decrease and elimination of blockade (1-11 h after dosage). Partial blockade and blockade of intermediate intensity induced concordant variations in RNA contents in the two cell types. This concordance in changes was impaired in conditions of complete blockade. Analysis of these results suggests that N-Ch synaptic processes in the sympathetic ganglion directly modulate quantitative changes in RNA levels in the neuron cytoplasm and represent a factor synchronizing the metabolism of neuronal and glial RNA.

[Lactate dehydrogenase isoenzymes in sympathetic neurons and satellite glial cells in normal animals and after nicotinic acetylcholine receptor blockade].

Using the method of integral cytophotometry, lactate dehydrogenase (LDH) N- and M-isoform activity was determined in neurons and satellite glial cells in tissue sections of rabbit cervical cranial sympathetic ganglion under normal conditions and after experimentally induced total and partial pharmacological blockade of nicotinic acetylcholine receptors (N-CR). In normal animals, activity of N- and M-isoforms was demonstrated in both cellular types, however, their LDH isoenzyme profile was different: N-isoforms prevailed in neurons, while M-isoforms were more active in glial cells. After partial and total blockade, activity of LDH, N- and M-isoforms was significantly reduced in neurons in proportion to the number of N-CR blocked. In satellite glial cells, the increase of blockade intensity was accompanied by the reduction of only M-isoform activity, while that of N-isoforms remained unchanged. After partial N-CR blockade, satellite glial cell LDH isoenzyme profile was shifted in the direction of a neuronal one, and after total blockade became indistinguishable from LDH isoenzyme profile of intact neurons. It is suggested that lactate formation in satellite glial cells is induced by N-cholinergic synapse activity, which directly mediates the neuron-glial interaction and participates in control of LDH enzyme system activity in sympathetic neurons.

[Adenylate level and lactate dehydrogenase (LDH) in the sympathetic ganglion: the effect of N-cholinergic blockade].

Activity of LDH isoenzymes and the level of ATP, ADP, and AMP were determined in the rabbit superior cervical sympathetic ganglion under conditions of experimentally induced partial or total blockade of N-cholinergic (N-CE) synapses. This blockade changed the spectrum of LDH isoenzymes: partial blockade was accompanied by disappearance of LDH-4 and LDH-5; total blockade also caused disappearance of LDH-3. LDH-1 and LDH-2 that remained in the isoenzyme spectrum as well as total LDH activity decreased significantly. Under conditions of total N-CE blockade total activity of LDH represented 9% of control, whereas activity of LDH-1 and LDH-2 represented 16% of control. ATP content decreased by 53 and 93% under conditions of partial and total N-CE blockade, respectively. The levels of ADP and AMP decreased by 33 and 65 and 80 and 56%, respectively. Results of the present study suggest that activity of N-CE synapses is a crucial factor involved into the energy homeostasis of the sympathetic ganglion.

[Participation of N-cholinergic synapses in the regulation of RNA metabolism of sympathetic neurons and satellite glial cells].

Cytoplasmic rRNA content in neuronal and in satellite glial cells of cervical sympathetic ganglion was defined using photographic cytophotometry during the pharmacological blockade of N-cholinergic (N-CE) synapses, induced by the injection of N-cholinolytic drug dimecoline (10, 30 and 50 mg/kg subcutaneously), as well as during the period of blockade gradual resolution and termination (1-11 hours after treatment with cholinolytic drug). The partial blockade and average intensity blockade induced coordinated oscillations of rRNA content in both types of cells. The coordination of these changes was violated during the complete blockade. Analysis of the results obtained suggested that N-CE synaptic processes directly modulated quantitative changes of rRNA in neuronal cytoplasm could be a factor synchronizing the metabolism of neuronal and glial rRNA in a sympathetic ganglion.

Possible involvement of lactate in neuroglial interaction through nicotinic cholinergic synapses in the cranial cervical sympathetic ganglion.

Activities of LDH and its H- and M-isoforms in neurons and satellite gliocytes of the cranial cervical sympathetic ganglion in rabbits under normal conditions and during nicotinic cholinergic synapse blockade were evaluated by integral cytophotometry in tissue sections. Normally activity of H-isoform predominates in neurons and M-isoform in satellite gliocytes. Blockade of the cranial cervical sympathetic ganglion significantly decreased LDH activity (H- and M-isoforms) in neurons in direct proportion to the number of blocked nicotinic cholinergic receptors. Activity of M-isoform in satellite gliocytes decreased with increasing the degree of blockade, while activity of H-isoform did not change. The isoenzyme profile of LDH in satellite gliocytes reached the level of intact neurons. Presumably, lactate production in satellite gliocytes is regulated by sympathetic neurons through nicotinic cholinergic synapses.

Isoenzyme profile of lactate dehydrogenase in the cranial cervical sympathetic ganglion under normal conditions and during synaptic blockade.

The isoenzyme profile of lactate dehydrogenase in the cranial cervical sympathetic ganglion of rabbits was studied under normal conditions and during blockade of nicotinic cholinergic synapses. Under normal conditions this profile was presented by 5 isoforms of the enzyme (lactate dehydrogenases 1, 2, 3, 4, and 5). Activity of H-isoforms prevailed. Blockade was accompanied by heterotropic allosteric inhibition of lactate dehydrogenase isoforms. H- and M-isoforms underwent simultaneous changes. Activity of H-isoforms sharply decreased. However, the ratio between lactate dehydrogenases 1 and 2 during complete or partial blockade did not differ from that observed in experiments with the intact ganglion. M-isoforms (lactate dehydrogenases 4 and 5) disappeared during partial blockade. Activity of hybrid lactate dehydrogenase 3 significantly decreased and was undetected during partial and complete blockade, respectively. Our results indicate that enzyme activity and isoenzyme profile of lactate dehydrogenase are determined by function of nicotinic synapses.

[Functional changes in the chromatophilic substance and RNA content of the cytoplasm of sympathetic neurons in the presence of synaptic transmission disorders]. / Funktsional'nye izmeneniia khromatofil'nogo veshchestva i soderzhaniia RNK v tsitoplazme simpaticheskikh neironov pri narushenii sinapticheskoi peredachi.

At a disturbed synaptic transmission in the rabbit cranial sympathetic cervical ganglion in histological sections stained with gallocyanin chrome alum, distribution of the chromatophilic substance was studied, and in the same sections RNA content was determined cytophotometrically. In ganglia of intact animals three groups of neurons with various structure of the chromatophilic substance were defined and their quantitative relation was stated. After administration of various doses of the ganglio-blockader, there was an essential difference in the changes of the chromatophilic substance, but they were unitypical in mono- and double-nuclear neurons. In the same cells, the synaptic blockade produced a rather great increase in the content of the cytoplasmic RNA. Comparing the quantitative data with the visual observation results, a conclusion was made that it is not reliable to use any changes in the chromatophilic substance as a criterium on quantitative shifts in the neuronal RNA and for the morphological test of the neuronal functional activity.

[Dynamics of the cytoplasmic RNA content in the sympathetic neurons in a synaptic block]. / Dinamika soderzhaniaa tsitoplazmaticheskoi RNK simpaticheskikh neironov pri sinapticheskoi blokade.

Cytophotometry of RNA was carried out in the neurons of the superior cervical sympathetic ganglion of the adult rabbits under the effect of various doses of the ganglion blocking agent dimecolin. Marked oscillations in the RNA content were observed in the neuronal cytoplasm after administration of the preparation. Dynamics of revealed shifts significantly differed depending on the dose of the preparation, and it was similar in the mono- and binucleated neurons. On the basis of the analysis of the character of variations revealed it is suggested that the synaptic processes have an important role in the development of quantitative RNA shifts in postsynaptic neurones. The results obtained indicate that the functional activity in some ganglioid neurones increases after administration of various doses of dimecolin during the blockade and after its termination.

[Effect of histological processing on the nucleic acid, free nucleotide and protein content in the cranial cervical sympathetic ganglion]. / Vliianie gistologicheskoi obrabotki na soderzhanie nukleinvykh kislot, svobodnykh nukleotidov i belka v kranial'nom sheinom simpaticheskom ganglii.

By means of biochemical techniques, the stability of free nucleotide contents has been demonstrated in the rabbit sympathetic node fixed in the Carnoy solution for 1--2 hours at 4 degrees C. A 2 hour fixation at room temperature and at 37 degrees, however, results in a considerable loss of free nucleotides--of the total amount in the fresh ganglion tissue, resp., 66.6 and 74.5%. According to these data, the Carnoy solution can be successfully used for a quantitative precipitation of free nucleotides. A histological treatment of this Carnoy fixed sympathetic ganglion (for 2 hours, at 4 degrees) causes no losses of nucleic acids, free nucleotides or protein, does not affect RNA, DNA or protein contents, but leads, however, to a remarkable loss of free nucleotides--up to 45%.

[Effect of fixation in Carnoy's solution on nucleic acid and protein concentration in the rabbit superior cervical sympathetic ganglion]. / Vliianie fiksatsii v shidkosti Karnua na soderzhanie nukleinovykh kislot i belka v verkhnem sheinom simpaticheskom ganglii krolika.

By means of biochemical techniques, the stability of RNA, DNA and protein contents has been demonstrated in the rabbit superior cervical sympathetic node fixed in Carnoy's solution for 2 hours at 4 degrees C. A 2 hour fixation in Carnoy's fluid at room temperature, results, however, in a considerable loss of RNA (21%). The loss of DNA and protein did not exceed 2% of the total amount of the fresh ganglion tissue. Fixation at a higher temperature (37%) increased the loss percentage protein and of nucleic acids: 3.1, 5.5 and 42%, for protein, DNA and RNA, resp.

[Use of the ORIM-1 microscope for determining the thickness of paraffin sections]. / Primenenie mikroskopa ORIM-1 dlia opredeleniia tolshchiny parafinovykh srezov

On measuring the thickness of paraffin sections with the help of the "OPIM-1" microscope the error for sections of optimal thickness 5--10 mkm was no more than 3%.