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Daily isoflurane exposure increases barbiturate insensitivity in medullary respiratory and cortical neurons via expression of ε-subunit containing GABA ARs.

Hengen KB, Nelson NR, Stang KM, Johnson SM, Smith SM, Watters JJ, Mitchell GS, Behan M - PLoS ONE (2015)

Bottom Line: The parameters governing GABAA receptor subtype expression patterns are not well understood, although significant shifts in subunit expression may support key physiological events.We hypothesized that this plasticity may be a compensatory response to a chronic increase in inhibitory tone caused by increased central neurosteroid levels.We hypothesize that increased inhibitory tone in the respiratory control network and cortex causes a compensatory increase in ε-subunit-containing GABAARs.

View Article: PubMed Central - PubMed

Affiliation: Neuroscience Training Program, University of Wisconsin, Madison, Madison, Wisconsin, United States of America.

ABSTRACT
The parameters governing GABAA receptor subtype expression patterns are not well understood, although significant shifts in subunit expression may support key physiological events. For example, the respiratory control network in pregnant rats becomes relatively insensitive to barbiturates due to increased expression of ε-subunit-containing GABAARs in the ventral respiratory column. We hypothesized that this plasticity may be a compensatory response to a chronic increase in inhibitory tone caused by increased central neurosteroid levels. Thus, we tested whether increased inhibitory tone was sufficient to induce ε-subunit upregulation on respiratory and cortical neurons in adult rats. Chronic intermittent increases in inhibitory tone in male and female rats was induced via daily 5-min exposures to 3% isoflurane. After 7d of treatment, phrenic burst frequency was less sensitive to barbiturate in isoflurane-treated male and female rats in vivo. Neurons in the ventral respiratory group and cortex were less sensitive to pentobarbital in vitro following 7d and 30d of intermittent isoflurane-exposure in both male and female rats. The pentobarbital insensitivity in 7d isoflurane-treated rats was reversible after another 7d. We hypothesize that increased inhibitory tone in the respiratory control network and cortex causes a compensatory increase in ε-subunit-containing GABAARs.

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Spontaneous neuronal activity resistance to bath-applied pentobarbital in medullary and cortical slices from 7d isoflurane-treated rats.Spontaneous activity of neurons in the region anatomically consistent with the VRC and PreBötC (all labeled as VRC neurons) are insensitive to bath-applied pentobarbital (300 μM) following 7d isoflurane treatment. A, Mean normalized firing rate of VRC (n = 62, open circles) and CTX (n = 37, filled circles) neurons from male rats in response to 300 μM pentobarbital (applied at vertical bar, t = 30 min). B, Pentobarbital had equivalent depressive effects on VRC (n = 37) and CTX (n = 47) neurons from virgin female rats. C, Following pentobarbital treatment, spontaneous activity of VRC neurons (n = 62) from male rats treated with intermittent isoflurane for 7d were significantly more active than VRC neurons from untreated control rats. The response of cortical neurons (n = 49) to pentobarbital was non-significantly shifted in the same direction. D, Likewise, in female rats treated with isoflurane for 7d, pentobarbital failed to inhibit spontaneous neuronal activity in the VRC neurons (n = 40). CTX neurons (n = 51) trended non-significantly towards an increase in activity compared to untreated control female rats. E, This effect was reversible, as 7d of recovery following 7d of intermittent isoflurane shifted spontaneous activity of VRC (n = 27) and CTX neurons (n = 38) in rats toward baseline values. F., 30d of intermittent isoflurane significantly attenuated the pentobarbital response in VRC (n = 40) and CTX neurons (n = 32) in male rats. To confirm the presence of pentobarbital-insensitive GABAARs, muscimol (20 μM) was applied at 90 min (dashed vertical line). Nearly all neurons that were resistant to pentobarbital were inhibited by muscimol. G, In female rats exposed to 30d isoflurane treatment, the response of VRC (n = 123) and CTX (n = 83) neurons to pentobarbital was significantly attenuated. These neurons were also silenced by muscimol (dashed vertical line). (H,I) The average steady-state responses to bath-applied pentobarbital are quantified for the different isoflurane treatments and recovery for male rats (C) and female rats (D). The asterisk indicates p<0.05 for comparison to control while the pound sign indicates p<0.05 for comparison to 7d treatment.
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pone.0119351.g002: Spontaneous neuronal activity resistance to bath-applied pentobarbital in medullary and cortical slices from 7d isoflurane-treated rats.Spontaneous activity of neurons in the region anatomically consistent with the VRC and PreBötC (all labeled as VRC neurons) are insensitive to bath-applied pentobarbital (300 μM) following 7d isoflurane treatment. A, Mean normalized firing rate of VRC (n = 62, open circles) and CTX (n = 37, filled circles) neurons from male rats in response to 300 μM pentobarbital (applied at vertical bar, t = 30 min). B, Pentobarbital had equivalent depressive effects on VRC (n = 37) and CTX (n = 47) neurons from virgin female rats. C, Following pentobarbital treatment, spontaneous activity of VRC neurons (n = 62) from male rats treated with intermittent isoflurane for 7d were significantly more active than VRC neurons from untreated control rats. The response of cortical neurons (n = 49) to pentobarbital was non-significantly shifted in the same direction. D, Likewise, in female rats treated with isoflurane for 7d, pentobarbital failed to inhibit spontaneous neuronal activity in the VRC neurons (n = 40). CTX neurons (n = 51) trended non-significantly towards an increase in activity compared to untreated control female rats. E, This effect was reversible, as 7d of recovery following 7d of intermittent isoflurane shifted spontaneous activity of VRC (n = 27) and CTX neurons (n = 38) in rats toward baseline values. F., 30d of intermittent isoflurane significantly attenuated the pentobarbital response in VRC (n = 40) and CTX neurons (n = 32) in male rats. To confirm the presence of pentobarbital-insensitive GABAARs, muscimol (20 μM) was applied at 90 min (dashed vertical line). Nearly all neurons that were resistant to pentobarbital were inhibited by muscimol. G, In female rats exposed to 30d isoflurane treatment, the response of VRC (n = 123) and CTX (n = 83) neurons to pentobarbital was significantly attenuated. These neurons were also silenced by muscimol (dashed vertical line). (H,I) The average steady-state responses to bath-applied pentobarbital are quantified for the different isoflurane treatments and recovery for male rats (C) and female rats (D). The asterisk indicates p<0.05 for comparison to control while the pound sign indicates p<0.05 for comparison to 7d treatment.

Mentions: Consistent with prior data [39, 40, 20], in slices from untreated control male (n = 6) and female rats (n = 6), 60 min exposure to bath-applied 300 μM pentobarbital inhibited spontaneous activity in VRC neurons to 44 ± 15% of baseline (n = 103 neurons, p<0.05) and cortical (CTX) neurons to 24 ± 8% of baseline (n = 84 neurons, p<0.05) compared to time controls (n = 5 animals, 56 VRC neurons and 48 CTX neurons, 136 ± 19% and 123 ± 19% of baseline, respectively) (Fig. 2A, B). There were no significant differences between untreated male and female animals.


Daily isoflurane exposure increases barbiturate insensitivity in medullary respiratory and cortical neurons via expression of ε-subunit containing GABA ARs.

Hengen KB, Nelson NR, Stang KM, Johnson SM, Smith SM, Watters JJ, Mitchell GS, Behan M - PLoS ONE (2015)

Spontaneous neuronal activity resistance to bath-applied pentobarbital in medullary and cortical slices from 7d isoflurane-treated rats.Spontaneous activity of neurons in the region anatomically consistent with the VRC and PreBötC (all labeled as VRC neurons) are insensitive to bath-applied pentobarbital (300 μM) following 7d isoflurane treatment. A, Mean normalized firing rate of VRC (n = 62, open circles) and CTX (n = 37, filled circles) neurons from male rats in response to 300 μM pentobarbital (applied at vertical bar, t = 30 min). B, Pentobarbital had equivalent depressive effects on VRC (n = 37) and CTX (n = 47) neurons from virgin female rats. C, Following pentobarbital treatment, spontaneous activity of VRC neurons (n = 62) from male rats treated with intermittent isoflurane for 7d were significantly more active than VRC neurons from untreated control rats. The response of cortical neurons (n = 49) to pentobarbital was non-significantly shifted in the same direction. D, Likewise, in female rats treated with isoflurane for 7d, pentobarbital failed to inhibit spontaneous neuronal activity in the VRC neurons (n = 40). CTX neurons (n = 51) trended non-significantly towards an increase in activity compared to untreated control female rats. E, This effect was reversible, as 7d of recovery following 7d of intermittent isoflurane shifted spontaneous activity of VRC (n = 27) and CTX neurons (n = 38) in rats toward baseline values. F., 30d of intermittent isoflurane significantly attenuated the pentobarbital response in VRC (n = 40) and CTX neurons (n = 32) in male rats. To confirm the presence of pentobarbital-insensitive GABAARs, muscimol (20 μM) was applied at 90 min (dashed vertical line). Nearly all neurons that were resistant to pentobarbital were inhibited by muscimol. G, In female rats exposed to 30d isoflurane treatment, the response of VRC (n = 123) and CTX (n = 83) neurons to pentobarbital was significantly attenuated. These neurons were also silenced by muscimol (dashed vertical line). (H,I) The average steady-state responses to bath-applied pentobarbital are quantified for the different isoflurane treatments and recovery for male rats (C) and female rats (D). The asterisk indicates p<0.05 for comparison to control while the pound sign indicates p<0.05 for comparison to 7d treatment.
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Related In: Results  -  Collection

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pone.0119351.g002: Spontaneous neuronal activity resistance to bath-applied pentobarbital in medullary and cortical slices from 7d isoflurane-treated rats.Spontaneous activity of neurons in the region anatomically consistent with the VRC and PreBötC (all labeled as VRC neurons) are insensitive to bath-applied pentobarbital (300 μM) following 7d isoflurane treatment. A, Mean normalized firing rate of VRC (n = 62, open circles) and CTX (n = 37, filled circles) neurons from male rats in response to 300 μM pentobarbital (applied at vertical bar, t = 30 min). B, Pentobarbital had equivalent depressive effects on VRC (n = 37) and CTX (n = 47) neurons from virgin female rats. C, Following pentobarbital treatment, spontaneous activity of VRC neurons (n = 62) from male rats treated with intermittent isoflurane for 7d were significantly more active than VRC neurons from untreated control rats. The response of cortical neurons (n = 49) to pentobarbital was non-significantly shifted in the same direction. D, Likewise, in female rats treated with isoflurane for 7d, pentobarbital failed to inhibit spontaneous neuronal activity in the VRC neurons (n = 40). CTX neurons (n = 51) trended non-significantly towards an increase in activity compared to untreated control female rats. E, This effect was reversible, as 7d of recovery following 7d of intermittent isoflurane shifted spontaneous activity of VRC (n = 27) and CTX neurons (n = 38) in rats toward baseline values. F., 30d of intermittent isoflurane significantly attenuated the pentobarbital response in VRC (n = 40) and CTX neurons (n = 32) in male rats. To confirm the presence of pentobarbital-insensitive GABAARs, muscimol (20 μM) was applied at 90 min (dashed vertical line). Nearly all neurons that were resistant to pentobarbital were inhibited by muscimol. G, In female rats exposed to 30d isoflurane treatment, the response of VRC (n = 123) and CTX (n = 83) neurons to pentobarbital was significantly attenuated. These neurons were also silenced by muscimol (dashed vertical line). (H,I) The average steady-state responses to bath-applied pentobarbital are quantified for the different isoflurane treatments and recovery for male rats (C) and female rats (D). The asterisk indicates p<0.05 for comparison to control while the pound sign indicates p<0.05 for comparison to 7d treatment.
Mentions: Consistent with prior data [39, 40, 20], in slices from untreated control male (n = 6) and female rats (n = 6), 60 min exposure to bath-applied 300 μM pentobarbital inhibited spontaneous activity in VRC neurons to 44 ± 15% of baseline (n = 103 neurons, p<0.05) and cortical (CTX) neurons to 24 ± 8% of baseline (n = 84 neurons, p<0.05) compared to time controls (n = 5 animals, 56 VRC neurons and 48 CTX neurons, 136 ± 19% and 123 ± 19% of baseline, respectively) (Fig. 2A, B). There were no significant differences between untreated male and female animals.

Bottom Line: The parameters governing GABAA receptor subtype expression patterns are not well understood, although significant shifts in subunit expression may support key physiological events.We hypothesized that this plasticity may be a compensatory response to a chronic increase in inhibitory tone caused by increased central neurosteroid levels.We hypothesize that increased inhibitory tone in the respiratory control network and cortex causes a compensatory increase in ε-subunit-containing GABAARs.

View Article: PubMed Central - PubMed

Affiliation: Neuroscience Training Program, University of Wisconsin, Madison, Madison, Wisconsin, United States of America.

ABSTRACT
The parameters governing GABAA receptor subtype expression patterns are not well understood, although significant shifts in subunit expression may support key physiological events. For example, the respiratory control network in pregnant rats becomes relatively insensitive to barbiturates due to increased expression of ε-subunit-containing GABAARs in the ventral respiratory column. We hypothesized that this plasticity may be a compensatory response to a chronic increase in inhibitory tone caused by increased central neurosteroid levels. Thus, we tested whether increased inhibitory tone was sufficient to induce ε-subunit upregulation on respiratory and cortical neurons in adult rats. Chronic intermittent increases in inhibitory tone in male and female rats was induced via daily 5-min exposures to 3% isoflurane. After 7d of treatment, phrenic burst frequency was less sensitive to barbiturate in isoflurane-treated male and female rats in vivo. Neurons in the ventral respiratory group and cortex were less sensitive to pentobarbital in vitro following 7d and 30d of intermittent isoflurane-exposure in both male and female rats. The pentobarbital insensitivity in 7d isoflurane-treated rats was reversible after another 7d. We hypothesize that increased inhibitory tone in the respiratory control network and cortex causes a compensatory increase in ε-subunit-containing GABAARs.

Show MeSH
Related in: MedlinePlus