85A. Škorjanc, S. Batagelj & K. Drašler: Thermal acclimatization does not affect the resting activity…
 ACTA BIOLOGICA SLOVENICA 
 LJUBLJANA 2007 Vol. 50, [t. 2: 85–92
Sprejeto (accepted): 2008-01-31
Thermal acclimatization does not affect the resting activity of type T1 
trichobothrium in the fi rebug (Pyrrhocoris apterus; Heteroptera)
Temperaturna aklimatizacija ne vpliva na mirovno aktivnost trihobotrija tipa T1 
pri rdečem škratcu (Pyrrhocoris apterus; Heteroptera)
Aleš ŠKORJANC, Samo BATAGELJ & Kazimir DRAŠLAR
Department of Biology, Biotechnical Faculty, University of Ljubljana, Večna pot 111,
SI-1000 Ljubljana, Slovenija; e-mail: ales.skorjanc@bf.uni-lj.si; kazimir.draslar@bf.uni-lj.si
Abstract. Firebugs (Pyrrhocoris apterus) pass through the winter in adult state. They 
undergo a series of physiological changes in order to increase their capacity to survive at low 
ambient temperatures. Nevertheless, even during winter their body temperature can rise up to 
28 °C for a few hours on a sunny day, which is comparable to summer conditions. To establish 
the impact of cold acclimatization on the function of mechanoreceptors, the resting activity of 
T1 type trichobothria warmed up to 20 °C is compared in cold and warm acclimatized animals, 
as well in animals acclimated to laboratory conditions. In cold acclimatized animals the mean 
resting activity is 3300 imp/min (SE 90, n=13), in warm acclimatized 3400 imp/min (SE 60, 
n=15), and in animals acclimated to laboratory conditions 3700 imp/min (SE 130, n=17). The 
similar trend is observed in the variability of inter-impulse time intervals. The mean coeffi cient 
of the interval variation is in both cold and warm acclimatized animals 0.28 (cold SE 0.013, 
n=13; warm SE 0.010, n=15), and in animals acclimated to laboratory conditions 0.33 (SE 0.012, 
n=17). These data show negligible differences between the three groups. We can conclude that 
the resting activity of type T1 trichobothrium remains limited to a narrow range, regardless of 
the phase of the acclimatization process. 
Keywords: Pyrrhocoris apterus, mechanoreception, trichobothrium, resting activity, thermal 
acclimatization.
Izvleček. Stenica, rdeči škratec (Pyrrhocoris apterus) preživlja hladno zimsko obdobje v 
stanju imaga. Preživetje v obdobju nizkih okoljskih temperatur ji omogoča niz fi zioloških pri-
lagoditev. Kljub temu se lahko v tem obdobju temperatura telesa ob sončnih dneh za kratek čas 
povzpne do 28 °C, kar približno ustreza poletnim pogojem. Vpliv aklimatizacije na delovanje 
čutilnih celic smo želeli ugotoviti s pomočjo mirovne aktivnosti trihobotrija tipa T1. Poskus je 
bil zasnovan tako, da smo telesno temperaturo, hladno, toplo in na temperaturo laboratorijskega 
okolja aklimatiziranih živali nastavili na 20°C in posneli spontano mirovno aktivnost. Izmerje-
ni povprečni vrednosti mirovne aktivnosti sta bili pri živalih aklimatiziranih na hladno 3300 
imp/min (SE 90, n=13) in pri živalih aklimatiziranih na toplo 3400 imp/min (SE 60, n=15). Pri 
živalih aklimatiziranih na laboratorijske pogoje pa je bila povprečna vrednost 3700 imp/min 
(SE 130, n=17). Podobno razmerje med skupinami se je pokazalo tudi pri analizi variabilnosti 
intervalov med živčnimi signali. Tako je bil pri hladno in toplo aklimatiziranih živalih povprečni 
koefi cient variacije intervalov 0.28 (hladno SE 0.013, n=13; toplo SE 0.010, n=15), pri živali 
aklimatiziranih na laboratorijsko okolje pa 0.33 (SE 0.012, n=17). Navedeni podatki kažejo, da 
so razlike med različno aklimatiziranimi živalmi neznatne. Torej je mirovna oziroma spontana 
aktivnost trihobotrija T1 proces, ki ne glede na stopnjo termične aklimatizacije, ostaja znotraj 
ozkega območja. 
86 Acta Biologica Slovenica, 50 (2), 2007
Ključne besede: Pyrrhocoris apterus, mehanorecepcija, trihobotrij, mirovna aktivnost, 
temperaturna aklimatizacija.
Introduction
Mechanosensitive fi liform sensillae in Heteroptera are commonly characterized by the name tri-
chobothria (TULLGREN 1918). As it is typical for fi liform sensillae, they have a simple structure with a 
single receptor neuron (reviewed by KEIL 1997). In the fi rebug (Pyrrhocoris apterus) they are classifi ed 
into three types. The registration of a single unit activity is very simple, therefore their functional 
properties have been well defi ned (DRAŠLAR 1973, 1980). For type T1 trichobothrium a constant resting 
activity in the level of 3000 imp/min is characteristic. Experiments in other insects have shown that the 
origin of the resting activity in fi liform sensilla is predominantly endogenous (BUNO et al. 1981; DAGAN 
& VOLMAN 1982; HAMON & GUILLET 1994; LANDOLFA & MILLER 1995). It has been suggested that the 
main source of the activity are depolarizing ion currents through the mechanosensitive transduction 
channels (BUNO et al. 1981; THURM 2001). The involvement of the transduction site in the resting 
activity of type T1 trichobothrium has been confi rmed also by our current investigations (DRAŠLAR & 
ŠKORJANC 1997; ŠKORJANC & DRAŠLAR 2001; paper in preparation). 
A typical feature of the fi rebug is its resistance to low temperatures. It can be seen active not 
only during the summer but also on the sunny winter days. Its cold hardiness depends on the ability 
to increase the supercooling capacity and therefore avoid freezing at temperatures below 0°C. The 
process of cold acclimatization has two phases. The fi rst is induced by the onset of the reproductive 
diapause in the late summer and the second by the lowering of the ambient temperatures in the late 
autumn. The acquired cold hardiness persists throughout the winter months, and is terminated by the 
rising temperatures in the early spring (SOCHA 1993; HODKOVA & HODEK 1994; KOŠTAL & ŠIMEK 2000; 
reviewed by HODKOVA & HODEK 2004). During the acclimatization the concentration of cryoprotective 
polyols in haemolymph is increased (KOŠTAL & ŠIMEK 2000; KOŠTAL et al. 2001, 2004; ŠLACHTA et al. 
2002) and the phospholipid membranes are remodelled (HODKOVA et al. 1999, 2002; TOMČALA et al. 
2006). Recent molecular studies have shown that the lipid composition of the cell membrane could 
affect the gating of mechanosensitive channels (KUNG 2005). The resting activity of type T1 tricho-
bothria could be therefore used as a probe for detecting seasonal changes in functional properties of 
the receptor due to the remodelling of the membrane lipids. On the basis of these presumptions we 
proposed a simple hypothesis. If the cell membranes are signifi cantly remodelled, the resting activity 
of preparations suddenly exposed to 20 °C should signifi cantly differ between cold and warm accli-
matized animals.
Material and Methods
Experiments were performed on the adult male fi rebugs (Pyrrhocoris apterus L., Hemipteroidea, 
Heteroptera). Animals were collected near Ljubljana. The animals, regarded as a cold acclimatized, were 
collected in January and February 2007, and the animals regarded as warm acclimatized in May and 
June 2007. Experiments were performed within a few weeks after the animals were collected. During 
this period they were maintained in glass containers, supplied with lime seeds and water ad libitum. 
To imitate natural conditions containers were kept outside. They were placed in a covered box, which 
was partly buried into the ground. The air temperature and the humidity were monitored by wireless 
weather station BAR 628 HG (Oregon Scientifi c USA). In the cold period the mean temperature was 
5°C (Tmin = 3.3 °C; Tmax = 8.1 °C) and in the warm period 17°C (Tmin = 13.5 °C; Tmax = 20.5 °C). The 
relative humidity was 90 ± 5 %. As a reference we used the data obtained between the years 2003 and 
87A. Škorjanc, S. Batagelj & K. Drašler: Thermal acclimatization does not affect the resting activity…
2007 in bugs that were kept in the laboratory at 20 ± 2 °C. These animals were collected on different 
locations and in different periods. We believe that this reference gave us a fair insight into the general 
level of the resting activity and therefore enabled a valid comparison of the resting activity in closely 
synchronized cold and warm acclimatized animals.
To avoid the thermal stress during preparation, cyanoacrylate adhesive (Loctite Super Attack, gel) 
was chosen instead of a warm wax-resin. Measurements showed that the glue has no toxic effects. 
Bugs were glued with their dorsal side down to a preparation holder. After the fi xation, their legs were 
removed and the wounds sealed with Kwik-Sil (WPI, Sarasota, USA). 
We recorded the resting activity of type T1 trichobothrium from the fi fth abdominal segment. For 
the nerve impulse registration tungsten electrodes were used (Goodfellow, Cambridge, UK). The re-
gistration electrode was inserted a few millimeters away from the trichobothrium, while the reference 
electrode was placed in the abdomen through the last abdominal segment (DRAŠLAR 1973, 1980). Signal 
was processed by an AI 402 preamplifi er and CyberAmp 320 signal conditioner (Molecular Devices, 
USA), and then digitalized by CED 1401 plus or CED 1401power interface (Cambridge Electronic 
Design, UK). The recording, spike sorting and analysis of the data were performed on a PC computer 
by Spike2 v.6.02 (Cambridge Electronic Design, UK) software. For statistical analysis Prism v.4.02 
(GraphPad Software Inc.) was used. 
The measurements were performed at the temperature of 20 ± 0.3°C. For this purpose the tempe-
rature of the preparation holder was regulated by a homemade controller based on a miniature Peltier 
element (RO3.3–3.9 Conrad). 
The frequency of the nerve impulses was a measure of the resting activity level. The relative 
variability of inter-impulse time intervals (IPI) was estimated with the coeffi cient of variation (CV). 
The coeffi cient of variation was determined as a ratio between IPI standard deviation and mean value. 
To compare the IPI distributions between the three groups of animals average IPI histograms were 
calculated. Only data obtained in trichobothria with long-term stability of the resting activity and 
normal response to a defl ection of the hair were considered as acceptable. In this paper data from 13 
cold and 15 warm acclimatized and 17 animals from the reference group are presented. 
In the fi eld experiments measurements of temperature were performed with K-type thermocouples 
(HH501DK, Omega). The body temperature was measured with the thermocouple inserted into the 
abdomen whereas the ambient temperature was determined with thermocouple placed next to the 
animal. The reference air temperature was measured in the shadow, 2 meters above the ground.
Results
The resting activity of type T1 trichobothrium was used as a probe for detecting the effect of the 
thermal acclimatization on the function of sensory cell. For the test temperature, at which the resting 
activity of cold and warm acclimatized animals was compared, we chose 20 °C. The data obtained 
throughout a period of four years in animals kept at the laboratory ambient temperature were used 
as reference. 
In the reference group the resting activity ranged from 2800 to 4600 imp/min (SD 540, n = 17), 
which was in a good agreement with published data (DRAŠLAR 1973, 1980). In the group of animals 
acclimatized to cold (winter) conditions the activity ranged from 2600 to 3700 imp/min (SD 300, n 
= 13). Similarly, in the group of animals acclimatized to warm (summer) conditions the range was 
between 2900 and 3800 imp/min (SD 230, n = 15) (Fig. 1a). The variability between individual pre-
parations was higher in the reference group. However, this is not surprising, considering the greater 
diversity in this group. The animals were of different origin, collected in different seasons over a longer 
period. The corresponding mean values of the resting activity rate were 3700 imp/min (SE 130) in 
the reference group, 3300 imp/min (SE 90) in the cold acclimatized and 3400 imp/min (SE 60) in the 
warm acclimatized group. No signifi cant difference was established in the rate between the cold and 
88 Acta Biologica Slovenica, 50 (2), 2007
warm acclimatized animals. This conclusion was confi rmed by the Student’s t-test (α= 0.05). The test 
also showed that the difference between the mean rate attained in the reference and the one attained in 
the cold acclimatized animals was statistically signifi cant (α = 0.05). However, the mean rates could 
not be distinguished between the reference and warm acclimatized animals. 
Figure 1: Resting activity in cold and warm acclimatized, and to laboratory conditions acclimated fi rebugs. A: 
Resting activity rate. B: Coeffi cient of variation of inter-impulse time intervals. Points represent individual 
experiments and lines the corresponding mean values. 
89A. Škorjanc, S. Batagelj & K. Drašler: Thermal acclimatization does not affect the resting activity…
Figure 2: Average inter-impulse time interval (IPI) histograms obtained in cold and warm acclimatized, and to 
laboratory conditions acclimated fi rebugs. 
Another parameter that describes the resting activity is the coeffi cient of variation (CV) of inter-
impulse intervals (IPI), which quantifi es the relative IPI variability and describes the regularity of 
impulse discharge. In the reference group the mean CV value was 0.33 (SE 0.01). In the cold as well 
as in the warm acclimatized group the CV was 0.28 (SE 0.01) (Fig. 1b). Again, no difference could 
be established between the cold and warm acclimatized animals and the CV was statistically higher in 
the reference group (α=0.05). However, even if these differences are considered, the resting activity 
rate and CV are limited to a very narrow range. This is well refl ected by the similarity of the average 
IPI histograms in the three compared groups of fi rebugs (Fig. 2). The process of thermal adaptation 
therefore has no obvious infl uence on the resting activity of T1 type trichobothrium.
To get an approximate estimation of the thermal conditions that allow the bug to become active, the 
temperature was measured in the natural habitat on a sunny winter day (19. 12. 2003). In the morning 
the bugs moved out from the shelter and assembled in the grooves of the tree bark. Despite the air 
temperature being close to 0°C, the temperature inside the grooves reached 25°C. However, the layer 
of the warm air extended only a few millimetres above the bark. The body temperature of the bug that 
was placed in the groove, exposed to the sunlight, increased from 5 °C to 28 °C. In comparison, in June 
(13. 6. 2003), at the air temperature of 31.5 °C, it increased to 37 °C (Fig. 3). Despite the difference of 
more than 30°C in the air temperature, the fi nal body temperature in December was only 10°C lower 
than in June. Although the fi rebugs spent most of the time during the winter at low temperatures, in 
the sun they were warmed up to summer-like temperatures for a few hours (Fig. 4).
90 Acta Biologica Slovenica, 50 (2), 2007
Figure 3: Warming of the fi rebug body. The temperature of the abdomen was measured. The animal with the body 
temperature of 5°C was placed into a bark groove exposed to the sun. The temperature of the air was 
measured in the shadow, 2 meters above the ground.
Figure 4: Daytime related change of the fi rebug body temperature. The position of the animal was in the bark 
groove on the sunny side of the tree. The measurement was done on 19. 12. 2003. 
91A. Škorjanc, S. Batagelj & K. Drašler: Thermal acclimatization does not affect the resting activity…
Discussion
In the summer a new generation of adult fi rebugs appears. They enter a diapause, and the animals 
that make it through the winter start breeding in the spring. To survive low temperatures during the win-
ter they undergo a series of processes, including a modifi cation of membrane phospholipids (HODKOVA 
et al. 1999, 2002; TOMČALA et al. 2006). Most of the data regarding the cold hardiness were obtained 
in the Bohemian population. The adaptation to low temperature can vary in geographically separated 
populations. The Bulgarian population, which is exposed to higher temperatures, for example, shows 
lower cold hardiness (KALUSHKOV & NEDVED 2000). Because the annual temperatures in Slovenia are 
similar to the Bohemian, similar adaptation can be expected. 
The composition of the cell membrane could affect the gating of the mechanosensitive ion channels 
(KUNG 2005), which are the most likely candidate for the source of the resting activity in fi liform sen-
silla (BUNO et al. 1981, THURM 2001). The resting activity of type T1 trichobothrium should therefore 
refl ect the effects of the acclimatization on the sensory cell function. 
Our results show no signifi cant difference neither in the rate nor the regularity of impulse discharge 
between the cold and warm acclimatized animals, when suddenly warmed to 20 °C. When interpreting 
these fi ndings, tissue specifi city has to be taken into consideration. Data regarding the membrane 
remodelling were obtained in the muscle and fat body (HODKOVA et al. 1999, 2002; TOMČALA et al. 
2006). Extending these fi ndings to the receptor cell seems appealing, however, it should be done with 
reservations. Perhaps the lipid composition of the sensory cell membranes is regulated in a way that 
ensures a constant resting activity throughout the year. Namely, the resting activity is important for the 
sensory receptor function, as it sets the reference level necessary for the stimulus encoding (DRAŠLAR 
1973, 1980). It is important that the sensory receptor remains calibrated when the animal gets active. 
In the sun the body temperature of the bug increases to summer-like temperature and remains high for 
a couple of hours even during the winter. Thus the body temperature, at which the fi rebugs are active, 
remains in the same range throughout the year. We suppose that the fi rebugs could have adapted in a 
way that enables them to survive low temperatures during the winter and also to preserve the sensitivity 
and calibration of the receptor when they get warmed and become active. 
Conclusion
The level of resting activity in trichobothria of type T1 in the fi rebug (Pyrrhocoris apterus) remains 
within narrow limits regardless of the season and thermal adaptation. 
Literature
BUNO W., MONTI-BLOCH L., MATEOS A., HANDLER P. 1981: Dynamic properties of cockroach »threadlike« 
hair sensilla. J. Neurobiol. 12, No. 2: 123–141.
DAGAN D., VOLMAN S. 1982: Sensory basis for directional wind detection in fi rst instar cockroaches, 
Periplaneta americana. J. Comp. Physiol. 147: 471–478. 
HAMON A., GUILLET JC. 1994: Some electrical properties of the cercal anemoreceptors of the cockroach, 
Periplaneta americana. Comp. Biochem. Physiol. 107A, No. 2: 357–368.
DRAŠLAR K. 1973: Functional properties of trichobothria in the bug Pyrrhocoris apterus (L.). J. Comp. 
Physiol. 84: 175–184.
DRAŠLAR K. 1980: Physiology of trichobothria in the bug Pyrrhocoris apterus (L.). Academia Scien-
tiarum et Artium Slovenica, Dissertationes XXII/5, p.p. 371–402.
DRAŠLAR K., ŠKORJANC A. 2003: Functional properties of trichobotria in the bug Pyrrhocoris apterus. 
In: ELSNER N. & H. ZIMMERMANN (eds.): The neurosciences from basic research to therapy. Stuttgart, 
Georg Thieme Verlag, p. 359.
92 Acta Biologica Slovenica, 50 (2), 2007
HODKOVA M., BERKOVA P., ZAHRADNIČKOVA H. 2002: Photoperiodic regulation of the phospholipid mo-
lecular species composition in thoracic muscles and fat body of Pyrrhocoris apterus (Heteroptera) 
via an endocrine gland, corpus allatum. J. Insect. Physiol. 48: 1009–1019.
HODKOVA M., HODEK I. 1994: Control of diapauses and supercooling by the retrocerebral complex in 
Pyrrhocoris apterus. Entomol. Exp. Appl. 70: 237–245.
HODKOVA M., HODEK I. 2004: Photoperiod, diapauses and cold-hardiness. Eur. J. Entomol. 101: 
445–458.
HODKOVA M., ŠIMEK P., ZAHRADNIČKOVA H., NOVAKOVA O. 1999: Seasonal changes in the phospholipid 
composition in thoracic muscles of a heteropteran, Pyrrhocoris apterus. Insect. Biochem. Mol. 
Biol. 29: 367–376.
KALUSHKOV P., NEDVED O. 2000: Cold hardiness of Pyrrhocoris apterus (Heteroptera: Pyrrhocoridae) 
from central and southern Europe. Eur. J. Entomol. 97: 149–153.
KEIL T. 1997: Functional morphology of insect mechanoreceptors. Microsc. Res. Tech. 39: 506–531. 
KOŠTAL V., ŠIMEK P. 2000: Overwintering strategy in Pyrrhocoris apterus (Heteroptera): the rela-
tions between life-cycle, chill tolerance and physiological adjustments. J. Insect. Physiol. 46: 
1321–1329.
KOŠTAL V., ŠLACHTA M., ŠIMEK P. 2001: Cryoprotective role of polyols independent of the increase in 
supercooling capacity in diapausing adults of Pyrrhocoris apterus (Heteroptera: Insecta). Comp. 
Biochem. Physiol. B. 130: 365–374.
KOŠTAL V., TOLLAROVA M., ŠULA J. 2004: Adjustments of the enzymatic complement for polyol bio-
synthesis and accumulation in diapausing cold-acclimated adults of Pyrrhocoris apterus. J. Insect. 
Physiol. 50: 303–313.
KUNG C. 2005: A possible unifying principle for mechanosensation. Nature 463: 647–654.
LANDOLFA M. A., MILLER J. P. 1995: Stimulus-response properties of cricket cercal fi liform receptors. 
J. Comp. Physiol. A. 177: 749–757.
SOCHA R. 1993: Pyrrhocoris apterus (Heteroptera) – an experimental model species: A review. Eur. 
J. Entomol. 90: 241–286. 
ŠKORJANC A., DRAŠLAR K. 2005: Spontaneous activity and adaptation mechanisms of trichobothria in 
the bug Pyrrhocoris apterus. Neuroforum (Heidelb.) 11, suppl.
ŠLACHTA M., BERKOVA P., VAMBERA J., KOŠTAL V. 2002: Physiology of cold-acclimation in non-diapau-
sing adults of Pyrrhocoris apterus (Heteroptera). Eur J Entomol 99: 181–187.
TOMČALA A., TOLLAROVA M., OVERGAARD J., ŠIMEK P., KOŠTAL V. 2006: Seasonal acquisition of chill 
tolerance and reconstruction of membrane glycerophospholipids in an overwintering insect: trig-
gering by low temperature, desiccation and diapause progression. J. Exp. Biol. 209: 4102–4114.
THURM U. 2001: Mechanosensorik. In: DUDEL J., MENZEL R. & R. F. SCHMIDT (eds.): Neurowissen-
schaften: Vom Molekül zur Kognition. Springer-Verlag, p. 339.
TULLGREN A. 1918: Zur Morphologie und Systematik der Hemipteren I. Entomol Tidskr., 38 (2): 
113–133.