Ž .Brain Research 796 1998 303–306

Short communication

Characterization of baroreceptor-related neurons in the monkey insular cortex

Zhi-Hua Zhang a, Patrick M. Dougherty b, Stephen M. Oppenheimer a,)

a Laboratory of Neurocardiology, CerebroÕascular DiÕision, Department of Neurology, The Johns Hopkins UniÕersity School of Medicine, Baltimore, MD,USA

b Department of Neurosurgery, The Johns Hopkins UniÕersity School of Medicine, Baltimore, MD, USA

Accepted 10 March 1998

Abstract

Insular neurons responsive to baroreceptor challenge have been identified in the rat, but not previously in primates. Characterization ofŽ .baroreceptor-related neurons was performed in 15 anesthetized monkeys Macaca fascicularis using extracellular single-unit recording

techniques. 131 units were investigated within the insula and surrounding regions. Based on their responses to phenylephrineŽ . Ž . Ž . Ž .hydrochloride PE and sodium nitroprusside SNP , three types of units were distinguished: 35r131 27% sympathoexcitatory SE ,

Ž . Ž . Ž . Ž .12r131 9% sympathoinhibitory SI and 84 64% null units. More baroreceptive units were found within the insula 38r73, 52% thanŽ . Ž .in surrounding areas 9r58, 16% p-0.001 . Lateralization was indicated with more baroreceptive units being encountered within the

Ž . Ž . Ž .right insula 28r44, 64% than the left 10r29, 34% ps0.02 . The majority of the responsive units were located within thedysgranular and granular insula in layers II, III and VrVI. These data suggest that cardiovascular representation may occur in the primateinsula as has been shown in other species. q 1998 Elsevier Science B.V. All rights reserved.

Keywords: Baroreceptor-related neuron; Extracellular recording; Insular cortex; Monkey

Viscerotopic organization has been demonstrated in thew xrat insular cortex 4,10,12,16 . Baroreceptor afferents con-

verge in the insula and changes in blood pressure, heartrate and production of cardiac arrhythmias have beengenerated on stimulation of the rat insula caudal to the

w xcrossing of the anterior commissure 10,12,16 . We re-w xcently characterized neurons in the rat insula 19 accord-

ing to their responses to the pressor agent phenylephrineŽ .hydrochloride PE and the depressor drug sodium nitro-

Ž . w xprusside SNP using criteria modified from Barman 3 .We have now explored the monkey insula to identifywhether similar cells can be demonstrated.

All procedures were approved by the Institutional Ani-mal Care and Use Committee of the Johns Hopkins Uni-versity and are consistent with the guideline of the NIHguide for the care and use of laboratory animals.

Ž .Fifteen adult male monkeys Macaca fascicularisŽweighing 4.5–7.5 kg were tranquillized with ketamine 10

) Corresponding author. Laboratory of Neurocardiology, Meyer 5-185,The Johns Hopkins Hospital, 600 North Wolfe Street, Baltimore, MD21287-7585. Fax: q1-410-614-9807.

.mgrkg, i.m. and catheterized for arterial pressure mea-surement and intravenous drug administration. Anesthesia

Ž .was induced with pentobarbital 25 mgrkg, i.v. and main-tained with an i.v. infusion of 5.0 mgrkgrh. Under deepanesthesia, the monkeys were paralyzed with pancuronium

Ž .bromide 0.1 mgrkgrh and artificially ventilated. End-tidal CO was kept at 3.5–4.5%. Core temperature was2

maintained at 37–388C with a Harvard homeothermic heat-ing pad.

The animals were fixed in a Transvertex stereotacticframe and the parietal bone drilled on both sides. ECGleads were placed subcutaneously in the lead II configura-tion. Extracellular recordings were made from sponta-neously firing neurons in the insular cortex and surround-

Žing regions with a glass microelectrode impedance 1–2.MV filled with a carbon filament. Coordinates for the

Žregions of study AP 11–20; ML 15–18; Depth 10–20 mm.from cortical surface were derived from the atlas of Szabo

w xand Cowan 13 adjusted for body weight. Single unitactivity, ECG, arterial pressure were displayed on an os-

Ž .cilloscope Hitachi VC 6155 and relayed to a computerrunning the BrainWave data acquisition and analysis pack-age. The acquisition software discriminated unit activity

0006-8993r98r$19.00 q 1998 Elsevier Science B.V. All rights reserved.Ž .PII S0006-8993 98 00268-6

( )Z.-H. Zhang et al.rBrain Research 796 1998 303–306304

from background and stored the data for off-line analysis.Spike size and configuration were continuously monitoredby the software to confirm that activity of the same cellwas recorded throughout the experiment.

Following unit isolation and stabilization for 2–3 min, 3min of spontaneous baseline activity was recorded. Then,arterial baroreceptors were challenged with i.v. administra-

Ž .tion of phenylephrine hydrochloride PE, 10 mgrkg orŽ .sodium nitroprusside SNP, 5 mgrkg in a volume of 0.2

ml 0.9% saline, followed by 0.2 ml 0.9% saline to flushthe dead space of the tubing. These doses induced "20–40mmHg change in arterial pressure. Responsive units werealso tested with a control injection of 0.2 ml 0.9% saline.Firing rate of the identified cell was determined over90–120 s after drug injection and compared with a similarpre-injection period. Changes in firing rate )"20% with-

out change in spike amplitude or shape were consideredw xsignificant 1,18,19 .

At the conclusion of each experiment, the bottom of thelast electrode track was marked by an electrolytic lesion.Cell sites were identified with respect to this referencepoint. The brain was removed and post-fixed in 10%formalin for at least 10 days. Frontal sections of 50 mmthickness were cut with a cryostatmicrotome and stainedwith 0.125% thionin solution. The slide was projectedusing a Zeiss Camera Lucida, and the recording sitesidentified with reference to a measured scale taking intoaccount the magnification, the distance between the record-ing site and the bottom of the microelectrode track. Correc-tion for brain shrinkage was made to ensure precise histol-ogy by measuring the depth of the track in the pre- andpost-fixed states.

Fig. 1. Firing patterns of baroreceptor-related and null units in the monkey insula. A: A SE unit showing cessation of firing with PE induced blood pressureŽ . Ž .rises Right and significantly increased firing with SNP induced blood pressure decreases Left . B: A SI unit showing firing rate increases after PE

Ž . Ž . Ž .injection Right and decreases following SNP administration Left . C: A non-responsive unit Null cell showing no significant change in firing rateŽ . Žfollowing blood pressure alteration. In each panel: Top trace shows the blood pressure BP ; bottom trace shows the histogram of neural firing spikesrs,

.binwidths1 s ; middle trace shows the summed neural spike waveforms showing consistency of amplitude and shape throughout the recording. Thearrows indicate drug administration. Time scales30 s.

( )Z.-H. Zhang et al.rBrain Research 796 1998 303–306 305

Table 1Distribution of baroreceptive units within and out IC

Right in IC Left in IC Right out IC Left out IC Total

SE Cell 21 9 3 2 35Si Cell 7 1 0 4 12Null Cell 16 19 22 27 84Total 44 29 25 33 131

ICs insular cortex; SEssympathoexcitatory; SIssympathoinhibitory.

Values are expressed as the mean"S.E.M. The resultswere assessed for intrainsular and extrainsular distributionand lateralization with Fisher’s exact test. Analysis ofpaired data used Student’s t-test; p-0.05 indicated statis-tical significance.

One hundred and thirty one units were investigated.Both right and left insular cortices were explored forbaroreceptor-related units. According to their response toPE and SNP, three types of units were categorized as

w xreported previously in the rat experiment 3,19 . Sympa-Ž .thoexcitatory SE units were identified by a significant

decreased firing rate following PE induced increase inblood pressure. The firing rate of these units increased in

Žtandem with SNP related reduction in blood pressure Fig.. Ž .1A . Sympathoinhibitory SI units increased firing rate

with PE injection and decreased their firing rate followingŽ .SNP administration Fig. 1B . Null units showed no re-

sponse to either SNP or PE induced changes in bloodŽ .pressure Fig. 1C .

The monkey insula was defined according to the cytoar-w xchitectonics and connectivity reported previously 2,8 .

Ž .About 47 of 131 36% units responded to PE and SNPinduced blood pressure alterations. More of these respon-

Ž .sive cells were located within the insula 38r73, 52%Ž . Žcompared with surrounding regions 9r58, 16% p-

. Ž .0.001 Table 1 . The distribution of these cells is summa-rized in Fig. 2. Lateralization was suggested with more

Žresponsive units being found within the right insula 28r44,. Ž . Ž64% than the left 10r29, 34% ps0.02, Fig. 2 and

. Ž .Table 1 . Also, 35 of 131 cells 27% showed sympathoex-Ž .citatory SE responses. The mean firing rates of these

units in response to PE and SNP are shown in Table 2. Thelatency of onset of SE responses to PE was 8.4"2.3 sŽ . Ž .range 0–18 s and 9.6"2.8 s 0–16 s for SNP. The

Ž .duration of the responses was 118"11 s 25–180 s forŽ .PE-related responses and 102"12 s 20–190 s for SNP

related responses. The latency and duration of the SI unit

Ž . Ž . Ž .Fig. 2. Distribution of SE v , SI ' and Null ` units within themonkey insular cortex and surrounding regions. Numbers indicate thedistance in mm anterior to the intraural zero point. AmygsAmygdala;CA sAnterior commissure; CdsDorsal cochlear nucleus; ChosOpticChiasm; ClsClaustrum; GpsGlobus pallidus; HypsHypothalamus;PutsPutamen; TOsOptic tract; VA s ventral anterior nucleus; VLmsVentral lateral nucleus of thalamus, medial part; VLosVentral lateralnucleus of the thalamus, oral part.

( )Z.-H. Zhang et al.rBrain Research 796 1998 303–306306

Table 2Ž .Effects of PE, SNP and NS on the Firing Rate spikesrs of SE, SI and

Null cells

Pre-PE Post-PE Pre-SNP Post-SNP Pre-NS Post-NS

SE cells 5.7"0.5 2.7"0.4) 4.0"0.4 6.8"1.3) 5.3"0.3 5.5"0.7SI cells 4.2"0.8 6.4"1.1) 6.0"1.0 3.5"1.4) 4.9"0.5 4.6"0.6Null cells 4.2"0.4 4.1"0.4 3.9"0.3 3.8"0.3 4.1"0.4 4.0"0.3

SEsSympathoexcitatory; SIsSympathoinhibitory; PEsPhenylephrine;SNPsSodium Nitroprusside; NSsNormal Saline. ) p-0.001.

Žresponses were similar to those of SE cells 9.6"1.8 sand 123.7"15.1 s for PE and 8.2"2.4 s and 113"16.7

. Ž .s for SNP, respectively . 84r131 64% null cells wereunresponsive to PE and SNP induced changes in bloodpressure. Fewer of these cells were encountered in the

Ž .insula 35r73, 48% than in the surrounding regionsŽ . Ž .49r58, 85% ps0.05, Table 1 . Saline injection pro-duced no significant changes in firing rate in any of these

Ž .neurons Table 1 . The majority of units responsive toŽ .changes in blood pressure SErSI were identified within

the dysgranular and granular insula in layers II, III andVrVI.

This study suggests that baroreceptor-related neuronsmay be identified within the monkey insula as has previ-

w xously been shown in the rat 5,19 . Likewise, under theseexperimental conditions, three distinct cell types can bedistinguished. In common with the rat, the predominant

w xresponse is of the SE type 19 . The suggested lateraliza-tion identified in this study can not be completely con-firmed because of the disparity between the sampling siteswithin the right and left insular cortices. Mesulam and

w xMufson 9 suggested from a consideration of connectivityŽstudies in the monkey that the anteroventral insula agranu-

.lar and anterior dysgranular regions may be involved inautonomic function. However, Augustine has concludedmore recently that this area may be primarily involved in

w x w xgustatoryralimentary function 2,17 as in the rat 15 . Theregion explored in this study abuts the autonomic zone as

w xoriginally described by Mesulam and Mufson 9 and alsoextends more caudally.

Insular involvement in cardiovascular control in themonkey has been suggested by earlier stimulation studiesw x6,14 . However, these lacked the precision of contempo-rary investigations due to the techniques available at thetime. Changes in heart rate and blood pressure were ob-tained on electrical stimulation of the anterior insula butlarge, constant voltages were used introducing concernsabout the specificity and localization of the response. Thisstudy however, supports these earlier observations andindicates the involvement of the insula in cardiovascularcontrol in the primate. Recent functional magnetic reso-nance imaging studies in humans have shown visceral

Ž .representation including baroreceptor input within thew xanterior ventrolateral insular cortex 7 . In addition, human

anterior insular stimulation may produce changes in bloodw xpressure and heart rate 11 . Consideration of all these

investigations indicates that the insula may be an importantsite of cardiovascular representation in primates includinghumans as has been previously shown in the rat.

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