Pollen germination and pollen tube growth are common to all seed plants, but these processes first developed in gymnosperms and still serve for their successful sexual reproduction. The main body of data on the reproductive physiology, however, was obtained on flowering plants, and one should be careful to extrapolate the discovered patterns to gymnosperms. In recent years, physiological studies of coniferous pollen have been increasing, and both the features of this group and the similarities with flowering plants have already been identified. The main part of the review is devoted to physiological studies carried out on conifer pollen. The main properties and diversity of pollen grains and pollination strategies in gymnosperms are described.

Pollen grain is a unique haploid organism characterized by two key physiological processes: activation of metabolism upon exiting dormancy and polar tube growth. In gymnosperms and flowering plants, these processes occur in different time frames and exhibit important features; identification of similarities and differences is still in the active phase. In angiosperms, the growth of male gametophyte is directed and controlled by its microenvironment, while in gymnosperms it is relatively autonomous. Recent reviews have detailed aspects of interaction between angiosperm female tissues and pollen such as interactions between peptides and their receptors; however, accumulated evidence suggests low-molecular communication, in particular, through ion exchange and ROS production, equally important for polar growth as well as for pollen germination. Recently, it became clear that ROS and ionic currents form a single regulatory module, since ROS production and the activity of ion transport systems are closely interrelated and form a feedback loop.

The concept of ROS as an important factor controlling pollen germination and tube growth has become generally accepted in the last decade. However, the relationship between various ROS and their significance for the success of in vivo germination and fertilization remained unexplored. For the present study, we collected Nicotiana tabacum stigma exudate on different stages of stigma maturity before and after pollination. Electron paramagnetic resonance (EPR) and colorimetric analysis were used to assess levels of O^•₂⁻ and H₂O₂ on stigma. Superoxide dismutase activity in the stigma tissues at each stage was evaluated zymographically. As the pistil matured, the level of both ROS decreased markedly, while the activity of SOD increased, and, starting from the second stage, the enzyme was represented by two isozymes: Fe SOD and Cu/Zn SOD, which was demonstrated by the in-gel inhibitory analysis. Selective suppression of Cu/Zn SOD activity shifted the ROS balance, which was confirmed by EPR. This shift markedly reduced the rate of pollen germination in vivo and the fertilization efficiency, which was estimated by the seed set. This result showed that hydrogen peroxide is a necessary component of stigma exudate, accelerates germination and ensures successful reproduction. A decrease in O^•₂⁻ production due to NADPH oxidase inhibition, although it slowed down germination, did not lead to a noticeable decrease in the seed set. Thus, the role of the superoxide radical can be characterized as less important.

Pollen germination in vivo on wet stigmas is assisted by the receptive fluid—stigma exudate. Its exact composition is still unknown because only some components have been studied. For the first time, hormonal screening was carried out, and the fatty acid (FA) composition of lipid-rich (Nicotiana tabacum) and sugar-rich (Lilium longiflorum) exudates was studied. Screening of exudate for the presence of plant hormones using HPLC-MS revealed abscisic acid (ABA) in tobacco stigma exudate at the two stages of development, at pre-maturity and in mature stigmas awaiting pollination, increasing at the fertile stage. To assess physiological significance of ABA on stigma, we tested the effect of this hormone in vitro. ABA concentration found in the exudate strongly stimulated the germination of tobacco pollen, a lower concentration had a weaker effect, increasing the concentration did not increase the effect. GC-MS analysis showed that both types of exudate are characterized by a predominance of saturated FAs. The lipids of tobacco stigma exudate contain significantly more myristic, oleic, and linoleic acids, resulting in a higher unsaturation index relative to lily stigma exudate lipids. The latter, in turn, contain more 14-hexadecenoic and arachidic acids. Both exudates were found to contain significant amounts of squalene. The possible involvement of saturated FAs, ABA, and squalene in various exudate functions, as well as their potential relationship on the stigma, is discussed.

Pollen is a highly specialized structure for sexual plant reproduction. Early stages of pollen germination require the transition from dormant state to active metabolism. In particular, an important role during this early phase of angiosperm pollen germination is played by H⁺-ATPase. Very little is known about pollen activation in gymnosperm species, and information on the involvement of H⁺-ATPase is lacking. We tracked four indicators characterizing the physiological state of pollen: membrane potential, intracellular pH, anion efflux and oxygen uptake, in order to monitor the dynamics of activation in Picea pungens. Based on pH dynamics during activation, we assumed the important role of H⁺-ATPase in spruce pollen germination. Indeed, germination was severely suppressed by P-type ATPase inhibitor orthovanadate. In spruce pollen tubes, a pronounced pH gradient with a maximum in the apical zone was found, which was different from the pollen tubes of flowering plants. Using orthovanadate and fusicoccin, we found that the proton pump is largely responsible for maintaining the gradient. Immunolocalization of the enzyme in pollen tubes showed that the distribution of H⁺-ATPase generally coincides with the shape of the pH gradient: its maximum accumulation is observed in the apical zone.

Pollen–stigma interactions have been studied extensively because they play an important role in sexual reproduction and crop yield. The vast majority of studies have focused on dry stigmas, which are typical of many model and agricultural plants; however, the data obtained are difficult to apply to plants with wet stigmas, such as tomato and tobacco. Pollen germination in this case occurs in a liquid, an exudate, which has a complex, species-specific composition. UPLC-ESI-MS-based hormone screening was carried out for six plant genera belonging to Solanaceae, Bromeliaceae, and Gesneriaceae families and revealed jasmonic acid (JA), abscisic acid (ABA) and/or jasmonoyl-L-isoleucine (IleJA) in stigma exudates of tobacco, tomato, and Streptocarpus sp. To assess the physiological significance of plant hormones in stigma exudate we tested their effect in vitro, finding that JA, IleJA, and MeJa significantly stimulated germination of tobacco pollen, with JA being most effective in accordance with its predominance in the stigma exudate; furthermore, ABA stimulated pollen germination in all tested species including bromeliads despite the lack of this hormone in their exudates. Both JA and ABA had an anti-oxidant effect on germinating pollen. Possible functions of hormones and ROS in exudate as well as ways of implementing the anti-oxidant effect of phytohormones are discussed.