Turtle Brainstem Research Articles

Isolated adult turtle brainstems exhibit central hypoxic chemosensitivity

During hypoxia, red-eared slider turtles increase ventilation and decrease episodic breathing, but whether these responses are due to central mechanisms is not known. To test this question, isolated adult turtle brainstems were exposed to 240 min of hypoxic solution (bath PO2 = 32.6 ± 1.2 mmHg) and spontaneous respiratory-related motor bursts (respiratory event) were recorded on hypoglossal nerve roots. During hypoxia, burst frequency increased during the first 15 min, and then decreased during the remaining 35–240 min of hypoxia. Burst amplitude was maintained for 120 min, but then decreased during the last 120 min. The number of bursts/respiratory event decreased within 30 min and remained decreased. Pretreatment with either prazosin (α1-adrenergic antagonist) or MDL7222 (5-HT3 antagonist) blocked the hypoxia-induced short-term increase and the longer duration decrease in burst frequency. MDL7222, but not prazosin, blocked the hypoxia-induced decrease in bursts/respiratory event. Thus, during bath hypoxia, isolated turtle brainstems continued to produce respiratory motor output, but the frequency and pattern were altered in a manner that required endogenous α1-adrenergic and serotonin 5-HT3 receptor activation. This is the first example of isolated reptile brainstems exhibiting central hypoxic chemosensitivity similar to other vertebrate species.

Respiratory neuron characterization reveals intrinsic bursting properties in isolated adult turtle brainstems (Trachemys scripta)

It is not known whether respiratory neurons with intrinsic bursting properties exist within ectothermic vertebrate respiratory control systems. Thus, isolated adult turtle brainstems spontaneously producing respiratory motor output were used to identify and classify respiratory neurons based on their firing pattern relative to hypoglossal (XII) nerve activity. Most respiratory neurons (183/212) had peak activity during the expiratory phase, while inspiratory, post-inspiratory, and novel pre-expiratory neurons were less common. During synaptic blockade conditions, ∼10% of respiratory neurons fired bursts of action potentials, with post-inspiratory cells (6/9) having the highest percentage of intrinsic burst properties. Most intrinsically bursting respiratory neurons were clustered at the level of the vagus (X) nerve root. Synaptic inhibition blockade caused seizure-like activity throughout the turtle brainstem, which shows that the turtle respiratory control system is not transformed into a network driven by intrinsically bursting respiratory neurons. We hypothesize that intrinsically bursting respiratory neurons are evolutionarily conserved and represent a potential rhythmogenic mechanism contributing to respiration in adult turtles.

Severe hypoxia in isolated adult turtle brainstems decreases respiratory motor burst clustering, frequency, and amplitude

Responses of isolated amphibian and mammalian brainstems or slices to hypoxia are well characterized, but how isolated reptilian brainstems respond to severe hypoxia is unknown. Thus, we exposed isolated adult turtle brainstems to 4.0 h of hypoxia (bath PO2=36 ± 2 mmHg) while recording respiratory‐related motor bursts on hypoglossal nerves. In brainstems producing episodic discharge (n=6; baseline bursts/episode > 1.75), bursts/episode decreased from 2.3 ± 0.2 to 1.3 ± 0.2 (P<0.05) and burst frequency decreased by 32% (P<0.05) after 90 min of hypoxia. Burst amplitude steadily decreased and reached 64% of baseline (P<0.05) after 3.5 h of hypoxia. Burst shape did not appear to be altered. In brainstems producing singlet discharge (n=6; baseline bursts/episode < 1.25), burst amplitude decreased to 57% of baseline (P<0.05) after 4.0 h of hypoxia while other variables were unchanged. The decrease in bursts/episode observed in vitro was similar to the hypoxia‐induced decrease in breaths/episode observed in vivo, which required 5‐HT3 receptor activation (unpublished observations). Thus, we hypothesize that the hypoxia‐induced decrease in bursts/episode also requires 5‐HT3 receptor activation. Supported by T32 HL07654.

Regulation of respiratory-related hypoglossal motor output by α1 adrenergic and serotonin 5-HT3 receptor activation in isolated adult turtle brainstems

The effects of brainstem α(1) adrenergic receptor activation on respiratory control in reptiles are poorly understood. Isolated adult turtle brainstems were exposed to phenylephrine (α(1) adrenergic agonist) and respiratory motor bursts were recorded on hypoglossal nerves. Phenylephrine acutely increased burst frequency, amplitude (low concentrations only), and regularity of the time interval between the start of respiratory events (single or clustered bursts), and decreased bursts/respiratory event. Burst frequency and timing changes persisted during a 2.0 h washout. Acute increases in burst frequency and amplitude were blocked by prazosin (α(1) adrenergic antagonist). Pretreatment with prazosin and tropisetron (5-HT(3) antagonist) blocked the increase in respiratory event regularity, but did not alter the decrease in bursts/respiratory event. Intermittent phenylephrine application (4 × 5.0 min separated by 20 min) did not produce long-lasting changes in burst frequency and amplitude, bursts/respiratory event, or respiratory event regularity. Thus, sustained α(1) adrenergic receptor activation in turtle brainstems produces acute and long-lasting changes in respiratory burst frequency and pattern.

5-HT3 receptor-dependent modulation of respiratory burst frequency, regularity, and episodicity in isolated adult turtle brainstems

To determine the role of central serotonin 5-HT(3) receptors in respiratory motor control, respiratory motor bursts were recorded from hypoglossal (XII) nerve rootlets on isolated adult turtle brainstems during bath-application of 5-HT(3) receptor agonists and antagonists. mCPBG and PBG (5-HT(3) receptor agonists) acutely increased XII burst frequency and regularity, and decreased bursts/episode. Tropisetron and MDL72222 (5-HT(3) antagonists) increased bursts/episode, suggesting endogenous 5-HT(3) receptor activation modulates burst timing in vitro. Tropisetron blocked all mCPBG effects, and the PBG-induced reduction in bursts/episode. Tropisetron application following mCPBG application did not reverse the long-lasting (2h) mCPBG-induced decrease in bursts/episode. We conclude that endogenous 5-HT(3) receptor activation regulates respiratory frequency, regularity, and episodicity in turtles and may induce a form of respiratory plasticity with the long-lasting changes in respiratory regularity.

Microfluidic chambers used to test whether episodic breathing is regulated by the pons in turtle brainstems

Our goal is to locate and characterize neurons controlling respiratory motor burst timing. Episodic breathing (clusters of fictive breaths separated by apneas) is expressed in isolated adult turtle brainstems and is regulated by serotonin 5‐HT3 receptor activation (decreases bursts/episode) and blockade (increases bursts/episode). To test whether pontine 5‐HT3 receptors regulate episodic breathing, microfluidic chambers constructed from silicone elastomer were used to apply drugs selectively to the pons. In time controls (n=2), baseline episodic discharge (1.9 bursts/episode) was unchanged after 2.0 h (1.8 bursts/episode). Application of a 5‐HT3 agonist to the pons (n=1) switched respiratory motor bursts from 1.5 bursts/episode (baseline) to singlet discharge (1.0 bursts/episode) within 5 min. 5‐HT3 antagonist application to the pons (n=1) increased baseline discharge from 1.0 to 1.4 bursts/episode after 40 min. These observations suggest that the pons plays an important role in regulating episodic breathing. Funded by NSF (IOB 0517302).

Excitatory and inhibitory effects of opioid agonists on respiratory motor output produced by isolated brainstems from adult turtles (Trachemys)

To determine how central opioid receptor activation alters turtle breathing, respiratory-related hypoglossal (XII) motor bursts were recorded from isolated adult turtle brainstems during 60 min bath applications of agonists for delta- (DOR), kappa- (KOR), or nociceptin/orphanin (NOR) receptors. DADLE (DOR agonist) abolished XII burst frequency at 0.3-0.5 microM. DPDPE (DOR agonist) increased frequency by 40-44% at 0.01-0.1 microM and decreased frequency by 88+/-8% at 1.0 microM. U-50488 and U-59693 (KOR agonists) decreased frequency by 65-68% at 100 and 50 microM, respectively. Orphanin (NOR agonist) decreased frequency by 31-51% at 1.0-2.0 microM during the first 30 min period. Orphanin (0.5 and 2.0 microM) increased bursts/episode. Although morphine (10 microM) abolished frequency in nearly all brainstems, subsequent co-application of phenylephrine (alpha(1)-adrenergic agonist, 20-100 microM) with morphine restored activity to 16-78% of baseline frequency. Thus, DOR, KOR, and NOR activation regulates frequency and NOR activation regulates episodicity, while alpha(1)-adrenergic receptor activation reverses opioid-induced respiratory depression in turtles.

Episodic respiratory motor output is decreased by 5‐HT3 receptor agonists and increased by 5‐HT3 receptor antagonists in isolated adult turtle brainstems

Episodic breathing is a respiratory pattern marked by episodes of several consecutive breaths that are separated by non‐ventilatory pauses. Episodic breathing occurs naturally in some ectothermic vertebrates, but the underlying mechanisms are not known. Approximately 56% of isolated turtle brainstems produce single bursts of respiratory motor output on hypoglossal nerves (bursts/episode < 1.25), while 25% of isolated turtle brainstems produce episodic respiratory bursts (bursts/episode > 1.75; n=171; Johnson & Creighton, Am J Physiol Regul Integr Comp Physiol, 2005). Bath‐application of 5‐HT3 agonists, such as phenylbiguanide (PBG; 20 µM; n=8) or 1‐(m‐Chlorophenyl)biguanide hydrochloride (mCPBG; 50 µM; n=6), onto isolated turtle brainstems switched episodic respiratory bursts (2.04 ± 0.06 bursts/episode) into singlet bursts (1.31 ± 0.09 bursts/episode) after 1.0 h (P<0.001). Bath‐application of 5‐HT3 antagonists, such as tropisetron (50 µM; n=5) and MDL‐72222 (50 µM; n=3) tended to switch singlet respiratory bursts (1.05 ± 0.03 bursts/episode) into episodic bursts (1.52 ± 0.06 bursts/episode) after 2.0 h (P>0.05). These data suggest that 5‐HT3 receptors in the brainstem are involved in regulating episodic breathing in turtles.

Respiratory pattern in midline-lesioned brainstems and hemibrainstems from adult turtles

Discrete midline lesions uncouple left and right respiratory motor output in mammals, but not in frogs and lampreys. To address this question in reptiles, isolated adult turtle brainstems were cut along the midline while recording respiratory motor output (bursts of action potentials) on left and right hypoglossal (XII) nerves. XII motor bursts were synchronized as long as a small portion of the midline was still intact. When turtle brainstems were completely cut along the midline and separated into hemibrainstems, XII motor bursts were produced that could be abolished by mu-opioid receptor (MOR) activation or exposure to high pH (7.80) solution. Also, 13/57 hemibrainstems expressed episodic discharge (>1.75bursts/episode). To test whether crossed connections were necessary to express a long-lasting increase in burst frequency (i.e., frequency plasticity), phenylbiguanide (PBG, 5-HT(3) receptor agonist, 20microM) was bath-applied to hemibrainstems. Although PBG significantly increased burst frequency by 0.43+/-0.10bursts/min after 60min, no frequency plasticity was observed because burst frequency returned to near baseline levels after a 2-h washout. Thus, crossed connections in turtle brainstems synchronize respiratory motor output and are not required for normal respiratory pattern formation, but are required for PBG-dependent frequency plasticity.

Distribution of metabolic activity (cytochrome oxidase) and immunoreactivity to calcium-binding proteins in the turtle brainstem auditory nuclei

Using histochemical and immunohistochemical techniques, distribution of activity of oxidative mitochondrial enzyme cytochrome oxidase (CO) and of immunoreactivity to calcium-binding proteins has been studied in spiral ganglion and auditory nuclei of brainstem in two turtle species. It has been shown that immunoreactivity to calbindin, parvalbumin, and calretinin in neurons and neuropil of nuclei of cochlear and superior olivary complexes, in nucleus of lateral lemniscus, and in spiral ganglion neurons coincides topographically with the high CO activity. The similarity of the studied metabolic and neurochemical characteristics of these auditory centers in reptiles, birds, and mammals indicates the existence of some common principles of their organization in amniotes in spite of phylogenetic differences and peculiarities of specialization of the auditory system in different species.

Respiratory‐related pacemaker neurons in isolated turtle brainstems in vitro

For adult vertebrates, it is not known whether respiratory‐related pacemaker neurons exist and play a critical role in rhythm generation. To address this question, brainstems from adult turtles were isolated in vitro and respiratory motor bursts were recorded on hypoglossal (XII) nerve roots. Extracellular recordings of respiratory‐related neurons were acquired using multichannel silicon electrodes. While bathed in artificial cerebrospinal fluid, neurons (n=87) exhibited phasic behavior in relation to the XII motor bursts. Synaptic transmission was blocked with antagonists for GABAA (bicuculline), glycine (strychnine), AMPA (CNQX), and NMDA (APV) receptors. During synaptic transmission blockade, 14/87 neurons continued to fire with rhythmic bursts of action potentials and were considered putative pacemaker neurons. When [K+] was increased to 6.0–9.0 mM during ongoing synaptic blockade, 9/10 pacemaker neurons increased burst frequency and action potential firing rates within the bursts. Application of a mu‐opioid agonist (DAMGO) decreased burst frequency in 3/4 pacemaker neurons. These results suggest that respiratory pacemaker neurons, which respond to increased [K+] and mu‐opioid receptor activation, exist in adult turtles. We hypothesize that these pacemaker neurons may be critical for rhythm generation.

Analysis of quantal size of voltage responses to retinal stimulation in the accessory optic system

In the intact vertebrate central nervous system, the quantal nature of synaptic transmission is difficult to measure because the postsynaptic sites may be distributed along a tortuous dendritic tree that cannot be readily clamped spatially to a uniform potential. Titrating the intact brain's extracellular concentration of calcium ions is also challenging because of its strong buffering mechanisms. In this study, using a whole brain with eye attached preparation, quantal neurotransmission was examined in the turtle brainstem in vitro, by recording from accessory optic system neurons that receive direct input from visually responsive retinal ganglion cells. Unitary EPSPs, evoked by microstimulation of a single ganglion cell, were measured during whole cell current-clamp recordings. In this preparation, the neurons exhibit direction-selectivity, despite the hypoxic conditions. Bath application of cadmium to reduce calcium influx also reduced evoked EPSP amplitudes to that of the spontaneous synaptic events. Statistical analyses indicated that these evoked response amplitudes could be well fitted to a Poisson distribution for most brainstem neurons. Therefore, the spontaneous miniature excitatory synaptic events of approximately 1 mV, as also observed during spike blockade of the retina [Kogo, N., Ariel, M., 1997. Membrane properties and monosynaptic retinal excitation of neurons in the turtle accessory optic system. Journal of Neurophysiology 78, 614–627], are likely responses to the neurotransmitter of single vesicles release by retinal axon terminals.

The effects of unilateral eighth nerve block on fictive VOR in the turtle

Multiunit activity during horizontal sinusoidal motion was recorded from pairs of oculomotor, trochlear, or abducens nerves of an in vitro turtle brainstem preparation that received inputs from intact semicircular canals. Responses of left oculomotor, right trochlear and right abducens nerves were approximately aligned with leftward head velocity, and that of the respective contralateral nerves were in-phase with rightward velocity. We examined the effect of sectioning or injecting lidocaine (1–2 μL of 0.5%) into the right vestibular nerve. Nerve block caused a striking phase shift in the evoked response of right oculomotor and left trochlear nerves, in which (rightward) control responses were replaced by a smaller-amplitude response to leftward table motion. Such “phase-reversed” responses were poorly defined in abducens nerve recordings. Frequency analysis demonstrated that this activity was advanced in phase relative to post-block responses of the respective contralateral nerves, which were in turn phase-advanced relative to pre-block controls. Phase differences were largest (∼10°) at low frequencies (∼0.1 Hz) and statistically absent at 1 Hz. The phase-reversed responses were further investigated by eliminating individual canal input from the left labyrinth following right nVIII block, which indicated that the activation of the vertical canal afferents is the source of this activity.

Bilateral processing of vestibular responses revealed by injecting lidocaine into the eighth cranial nerve in vitro

Extracellular unit responses were recorded from the vestibular nucleus (VN) and medial longitudinal fasciculus during horizontal head rotation of an in vitro turtle brainstem in which the temporal bones remained attached. Units were characterized as type I or type II based on the responses to ipsiversive or contraversive rotation, respectively. Lidocaine injections (0.5–2 μl of 0.5%) into the root of the eighth cranial nerve within the cranium caused rapid effects on unit responses to head rotation. Responses of type I units were reduced by ipsilateral injection but enhanced following contralateral injection. On the other hand, type II units had their responses increased by ipsilateral injections yet decreased by contralateral injections. In approximately half of the type II cells, decrease of the contraversive response was accompanied by the appearance of latent ipsiversive activity. Our findings not only confirm that each eighth nerve has afferents that drive ipsiversive excitation of both vestibular nuclei but also suggest that both nerves compete to dominate a central neuron's vestibular response. These results may be inconsistent with the push–pull vestibular model in which each nerve drives the central neuron with a complementary response that enhances the vestibular output. An alternate model is described in which vestibular neurons receive bilateral excitation, and that excitatory input is antagonized by crossed inhibition during contraversive motion.

Abducens conditioning in in vitro turtle brain stem without cerebellum requires NMDA receptors and involves upregulation of GluR4-containing AMPA receptors.

Previous work showed that in vitro abducens eyeblink classical conditioning of turtle brain stem-cerebellum preparations involved NMDA-mediated mechanisms and redistribution of GluR4-containing AMPA receptors in the abducens motor nuclei. Since conditioning can be obtained in brain stem preparations without the cerebellum, we examined whether similar mechanisms were involved during conditioning of the brain stem alone. The results showed that conditioning could not be induced in the presence of the NMDA receptor antagonist dl-2-amino-5-phosphonovaleric acid (AP-5) and that abducens nerve conditioned responses, once initiated in normal saline, were significantly attenuated in the presence of AP-5. The effects of AP-5 did not generally depress physiological responsiveness of preparations because some abducens nerve reflexes were not significantly reduced by the compound. GluR4-containing AMPA receptors in the abducens motor nuclei were significantly upregulated and positively correlated with the levels of conditioning similar to that of preparations having an intact cerebellum. Furthermore, increased GluR4 subunits after brain stem conditioning was confirmed by Western blot analysis. These results suggest that NMDA receptor-mediated mechanisms and GluR4 upregulation may mediate in vitro abducens eyeblink classical conditioning and that these mechanisms reside in the brain stem eyeblink circuitry.

Synaptic pharmacology in the turtle accessory optic system.

The accessory optic system of the turtle (the basal optic nucleus, BON) receives both excitatory and inhibitory inputs that are direction-sensitive. When the dorsal midbrain is ablated, only the monosynaptic direction-sensitive input from the retina to the BON remains. To better understand the central visual processing performed by the accessory optic system, this study identifies the neurotransmitters and their receptors that mediate the synaptic excitation and inhibition of BON cells. We used a reduced in vitro turtle brainstem preparation in which the two eyes and brain were isolated pharmacologically. Patch recordings were made on BON neurons while drugs were applied to the brain, with the eyes bathed in control media and either exposed to visual pattern motion or subjected to electrical stimulation. An antagonist of the AMPA (alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid) subtype of glutamate receptor applied within the brain chamber blocked the visual responses. In response to electrical stimulation both excitatory and inhibitory synaptic events were blocked in BON cells, presumably by blocking direct excitation by retinal ganglion cell axons in the BON and indirect excitation of inhibitory interneurons elsewhere in the brainstem. An NMDA receptor antagonist was ineffective, even when the response was measured in a BON cell depolarized in Mg(2+)-free media. A GABA(A) receptor on the BON cell mediates the inhibitory responses to retinal stimulation. Injection of lidocaine into the contralateral eye caused an increase in spontaneous inhibitory post-synaptic potentials (IPSPs), suggesting that a tonic retinal output exists that reduces brainstem inhibition of BON cells. Also, there may be tonic inhibition of an excitatory path to BON neurons from within the brainstem, because bicuculline increased spontaneous excitatory post-synaptic potentials (EPSPs) observed in a BON cell without retinal input. These results indicate that the BON is a site of complex visual processing of competing visual signals and provide insight into how an interaction of excitation and inhibition creates a retinal slip signal in the accessory optic system.

Immunocytochemical localization of glutamate receptor subunits in the brain stem and cerebellum of the turtleChrysemys picta

The regional distribution of ionotropic (AMPA and NMDA) and metabotropic (mGluR1alpha) glutamate receptor subunits was examined in the brain stem and cerebellum of the pond turtle, Chrysemys picta, by using immunocytochemistry and light microscopy. Subunit-specific antibodies that recognize NMDAR1, GluR1, GluR4, and mGluR1alpha were used to identify immunoreactive nuclei in the brain stem and cerebellum. Considerable immunoreactivity in the turtle brain stem and cerebellum was observed with regional differences occurring primarily in the intensity of staining with the antibodies. The red nucleus, lateral reticular nucleus and cerebellum labeled intensely for NMDAR1 and moderately for GluR1. The cerebellum also labeled strongly for mGluR1alpha. All of the cranial nerve nuclei labeled intensely for NMDAR1 and to varying degrees for GluR1, GluR4, and mGluR1alpha. Counterstaining revealed the presence of neuronal somata where there were no immunoreactive neurons in individual nuclei. This finding suggests that there are subpopulations of immunoreactive neurons within a given nucleus that bear different glutamate receptor subunit compositions. The results suggest that the glutamate receptor subunit distribution in the brain stem and cerebellum of turtles is similar to that reported for rats. Additionally, there is considerable colocalization of NMDA and AMPA receptors as revealed by light microscopy. These results have implications for the organization of neural circuits that control motor behavior in turtles, and, generally, for the function of brain stem and cerebellar neural circuits in vertebrates.

Relationship between modulation of the cerebellorubrospinal system in the in vitro turtle brain and changes in motor behavior in rats: effects of novel sigma ligands

Saturation and competition binding studies showed that the turtle brain contains σ sites labeled by both [ 3H]di-o-tolylguanidine (DTG) and [ 3H](+)-pentazocine. There was a significant correlation between the IC 50 values of σ ligands for [ 3H]DTG sites in the turtle vs. rat brain, suggesting that the sites are comparable in the two species. In contrast, [ 3H](+)-pentazocine, which primarily labels σ1 sites in the rodent brain, labels a heterogeneity of sites in the turtle brain. In extracellular recordings from the in vitro turtle brainstem, some σ ligands enhanced the burst responses of red nucleus (RN) neurons (DTG, haloperidol, BD1031, BD1052, BD1069) while other σ ligands decreased the burst responses (BD1047, BD1063). Control compounds (turtle Ringer vehicle control, opiate antagonist naloxone, atypical neuroleptic sulpiride) had no significant effects on the RN burst responses recorded from the in vitro turtle brain. The ED 50s of the ligands for altering the burst responses in RN neurons from the turtle brain were correlated with their IC 50s for turtle brain sites labeled with [ 3H]DTG, but not [ 3H](+)-pentazocine; this pattern is identical to that previously reported in rats, where there is a correlation between the potencies of σ ligands for producing dystonic postures after microinjection into the rat RN and their binding to rat brain sites labeled with [ 3H]DTG, but not [ 3H](+)-pentazocine. When the novel σ ligands were microinjected into the rat RN, dystonic postures were produced by ligands that increased the burst duration of RN neurons in the turtle brain. Novel σ ligands that reduced the burst responses in the in vitro turtle brain have previously been reported to have no effects on their own when microinjected into the rat RN, but to block the dystonic postures produced by other σ ligands. Taken together, the data suggest that the opposite effects of the novel ligands in the turtle electrophysiological studies represent the actions of agonists vs. antagonists, and that the directionality of the effects has predictive value for the expected motor effects of the drugs.

Catecholaminergic modulation of respiratory rhythm in an in vitro turtle brain stem preparation.

An in vitro brain stem preparation from adult turtles was used to determine effects of dopamine (DA) and norepinephrine (NE) on the pattern of respiratory motor output recorded from hypoglossal nerve roots (XII). Bath-applied DA (10-200 microM) increased the frequency of respiratory bursts (peaks) from 0.9 +/- 0.2 to 2.4 +/- 0.3 (SE) peaks/min, resulting in a 99 +/- 9% increase in neural minute activity. R[+]-SCH-23390 (10 microM, D1 antagonist) and eticlopride (20 microM, D2 antagonist) attenuated the DA-mediated increase in peak frequency by 52 and 59%, respectively. On the other hand, the DA-receptor agonists apomorphine (D1, D2), quinelorane (D2), and SKF-38393 (D1) had no effect on peak frequency. Prazosin, an alpha1-adrenergic antagonist (250 nM) abolished the DA-mediated frequency increase. Although NE (10-200 microM) and phenylephrine (10-200 microM, alpha1-adrenergic agonist) increased peak frequency from 0.5 +/- 0.1 to 1.2 +/- 0.3 peaks/min and from 0.6 +/- 0.1 to 1. 0 +/- 0.2 peaks/min, respectively, these effects were not as large as that with DA alone. The data suggest that both dopaminergic and adrenergic receptor activation in the brain stem increase respiratory frequency in turtles, but the DA receptor-mediated increase is dependent on coactivation of alpha1-adrenergic receptors.

Hypoxia, temperature, and pH/CO2effects on respiratory discharge from a turtle brain stem preparation

An in vitro brain stem preparation from adult turtles (Chrysemys picta) was used to examine the effects of anoxia and increased temperature and pH/CO2 on respiration-related motor output. At pH approximately 7.45, hypoglossal (XII) nerve roots produced patterns of rhythmic bursts (peaks) of discharge (O.74 +/- 0.07 peaks/min 10.0 +/- 0.6 s duration) that were quantitatively similar to literature reports of respiratory activity in conscious, vagotomized turtles. Respiratory discharge was stable for 6 h at 22 degrees C; at 32 degrees C, peak amplitude and frequency progressively and reversibly decreased with time. Two hours of hypoxia had no effect on respiratory discharge. Acutely increasing bath temperature from 22 to 32 degrees C decreased episode and peak duration and increased peak frequency. Changes in pH/CO2 increased peak frequency from zero at pH 8.00-8.10 to maxima of 0.81 +/- 0.01 and 1.44 +/- 0.02 peaks/min at 22 degrees C (pH 7.32) and 32 degrees C (pH 7.46), respectively; pH/CO2 sensitivity was similar at both temperatures. We conclude that 1) insensitivity to hypoxia indicates that rhythmic discharge does not reflect gasping behavior, 2) increased temperature alters respiratory discharge, and 3) central pH/CO2 sensitivity is unaffected by temperature in this preparation (i.e., Q10 approximately 1.0).